WO2017104567A1 - Agent for stabilizing fibroblast growth factor 23 - Google Patents
Agent for stabilizing fibroblast growth factor 23 Download PDFInfo
- Publication number
- WO2017104567A1 WO2017104567A1 PCT/JP2016/086710 JP2016086710W WO2017104567A1 WO 2017104567 A1 WO2017104567 A1 WO 2017104567A1 JP 2016086710 W JP2016086710 W JP 2016086710W WO 2017104567 A1 WO2017104567 A1 WO 2017104567A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- fgf
- polyoxyethylene
- container
- nonionic surfactant
- solution
- Prior art date
Links
- 108090000569 Fibroblast Growth Factor-23 Proteins 0.000 title claims abstract description 311
- 102000004042 Fibroblast Growth Factor-23 Human genes 0.000 title claims abstract description 12
- 230000000087 stabilizing effect Effects 0.000 title abstract description 35
- 239000003795 chemical substances by application Substances 0.000 title abstract 4
- -1 polyoxyethylene Polymers 0.000 claims abstract description 191
- 238000000034 method Methods 0.000 claims abstract description 145
- 229920003171 Poly (ethylene oxide) Polymers 0.000 claims abstract description 127
- 239000002736 nonionic surfactant Substances 0.000 claims abstract description 82
- 239000012736 aqueous medium Substances 0.000 claims abstract description 45
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract description 25
- 239000004480 active ingredient Substances 0.000 claims abstract description 4
- 108010088751 Albumins Proteins 0.000 claims description 53
- 102000009027 Albumins Human genes 0.000 claims description 53
- 230000006641 stabilisation Effects 0.000 claims description 36
- 238000011105 stabilization Methods 0.000 claims description 36
- 239000003381 stabilizer Substances 0.000 claims description 30
- 108091003079 Bovine Serum Albumin Proteins 0.000 claims description 18
- 229940098773 bovine serum albumin Drugs 0.000 claims description 18
- 150000005215 alkyl ethers Chemical class 0.000 claims description 16
- 229920001214 Polysorbate 60 Polymers 0.000 claims description 14
- 229920003023 plastic Polymers 0.000 claims description 14
- 239000004033 plastic Substances 0.000 claims description 14
- 235000014113 dietary fatty acids Nutrition 0.000 claims description 13
- RTZKZFJDLAIYFH-UHFFFAOYSA-N ether Substances CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 13
- 239000000194 fatty acid Substances 0.000 claims description 13
- 229930195729 fatty acid Natural products 0.000 claims description 13
- 150000003973 alkyl amines Chemical class 0.000 claims description 11
- 239000004743 Polypropylene Substances 0.000 claims description 9
- 229920001155 polypropylene Polymers 0.000 claims description 9
- 239000004698 Polyethylene Substances 0.000 claims description 6
- 239000004793 Polystyrene Substances 0.000 claims description 6
- 229920000573 polyethylene Polymers 0.000 claims description 6
- 229920002223 polystyrene Polymers 0.000 claims description 6
- 238000004321 preservation Methods 0.000 claims description 3
- 241000894007 species Species 0.000 claims 1
- 102100024802 Fibroblast growth factor 23 Human genes 0.000 abstract description 309
- 101001051973 Homo sapiens Fibroblast growth factor 23 Proteins 0.000 abstract description 12
- 239000000243 solution Substances 0.000 description 116
- 239000000523 sample Substances 0.000 description 53
- 238000002360 preparation method Methods 0.000 description 37
- 238000005259 measurement Methods 0.000 description 32
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 26
- 125000000217 alkyl group Chemical group 0.000 description 22
- 235000002639 sodium chloride Nutrition 0.000 description 21
- 125000004432 carbon atom Chemical group C* 0.000 description 18
- YBJHBAHKTGYVGT-ZKWXMUAHSA-N (+)-Biotin Chemical compound N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 YBJHBAHKTGYVGT-ZKWXMUAHSA-N 0.000 description 16
- 241000047703 Nonion Species 0.000 description 14
- 239000000427 antigen Substances 0.000 description 13
- 102000036639 antigens Human genes 0.000 description 13
- 108091007433 antigens Proteins 0.000 description 13
- 238000010790 dilution Methods 0.000 description 13
- 239000012895 dilution Substances 0.000 description 13
- 239000011780 sodium chloride Substances 0.000 description 13
- 102000002260 Alkaline Phosphatase Human genes 0.000 description 12
- 108020004774 Alkaline Phosphatase Proteins 0.000 description 12
- 125000003438 dodecyl group Chemical group [H]C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])* 0.000 description 12
- 125000001421 myristyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 12
- 125000000913 palmityl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 12
- 125000004079 stearyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 12
- 239000006249 magnetic particle Substances 0.000 description 11
- 239000000203 mixture Substances 0.000 description 11
- 239000007864 aqueous solution Substances 0.000 description 9
- 235000010482 polyoxyethylene sorbitan monooleate Nutrition 0.000 description 9
- 229920000053 polysorbate 80 Polymers 0.000 description 9
- 238000011088 calibration curve Methods 0.000 description 8
- 239000002738 chelating agent Substances 0.000 description 8
- 238000000691 measurement method Methods 0.000 description 8
- 239000008363 phosphate buffer Substances 0.000 description 8
- 239000000244 polyoxyethylene sorbitan monooleate Substances 0.000 description 8
- 239000003755 preservative agent Substances 0.000 description 8
- 150000003839 salts Chemical class 0.000 description 8
- 102000008394 Immunoglobulin Fragments Human genes 0.000 description 7
- 108010021625 Immunoglobulin Fragments Proteins 0.000 description 7
- 229920001213 Polysorbate 20 Polymers 0.000 description 7
- 229960002685 biotin Drugs 0.000 description 7
- 235000020958 biotin Nutrition 0.000 description 7
- 239000011616 biotin Substances 0.000 description 7
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 7
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 7
- 239000012086 standard solution Substances 0.000 description 7
- SNRUBQQJIBEYMU-UHFFFAOYSA-N Dodecane Natural products CCCCCCCCCCCC SNRUBQQJIBEYMU-UHFFFAOYSA-N 0.000 description 6
- 150000001413 amino acids Chemical class 0.000 description 6
- 125000001204 arachidyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 6
- 125000002511 behenyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 6
- 239000012472 biological sample Substances 0.000 description 6
- 125000002704 decyl group Chemical group [H]C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])* 0.000 description 6
- 125000002960 margaryl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 6
- 125000001196 nonadecyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 6
- 125000001117 oleyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])/C([H])=C([H])\C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 6
- 125000002958 pentadecyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 6
- 125000002889 tridecyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 6
- 125000002948 undecyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 6
- SXGZJKUKBWWHRA-UHFFFAOYSA-N 2-(N-morpholiniumyl)ethanesulfonate Chemical compound [O-]S(=O)(=O)CC[NH+]1CCOCC1 SXGZJKUKBWWHRA-UHFFFAOYSA-N 0.000 description 5
- 102000018233 Fibroblast Growth Factor Human genes 0.000 description 5
- 108050007372 Fibroblast Growth Factor Proteins 0.000 description 5
- 229940126864 fibroblast growth factor Drugs 0.000 description 5
- 230000001900 immune effect Effects 0.000 description 5
- 238000004020 luminiscence type Methods 0.000 description 5
- 229920002114 octoxynol-9 Polymers 0.000 description 5
- 229920000259 polyoxyethylene lauryl ether Polymers 0.000 description 5
- 239000000725 suspension Substances 0.000 description 5
- FFJCNSLCJOQHKM-CLFAGFIQSA-N (z)-1-[(z)-octadec-9-enoxy]octadec-9-ene Chemical compound CCCCCCCC\C=C/CCCCCCCCOCCCCCCCC\C=C/CCCCCCCC FFJCNSLCJOQHKM-CLFAGFIQSA-N 0.000 description 4
- 125000005037 alkyl phenyl group Chemical group 0.000 description 4
- 239000000872 buffer Substances 0.000 description 4
- POULHZVOKOAJMA-UHFFFAOYSA-N dodecanoic acid Chemical compound CCCCCCCCCCCC(O)=O POULHZVOKOAJMA-UHFFFAOYSA-N 0.000 description 4
- IPCSVZSSVZVIGE-UHFFFAOYSA-N hexadecanoic acid Chemical compound CCCCCCCCCCCCCCCC(O)=O IPCSVZSSVZVIGE-UHFFFAOYSA-N 0.000 description 4
- 239000007788 liquid Substances 0.000 description 4
- 125000001400 nonyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 4
- 125000002347 octyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 4
- 239000003921 oil Substances 0.000 description 4
- 229920002503 polyoxyethylene-polyoxypropylene Polymers 0.000 description 4
- 229940068968 polysorbate 80 Drugs 0.000 description 4
- 108090000623 proteins and genes Proteins 0.000 description 4
- 229920006395 saturated elastomer Polymers 0.000 description 4
- 150000004671 saturated fatty acids Chemical class 0.000 description 4
- 235000021122 unsaturated fatty acids Nutrition 0.000 description 4
- 150000004670 unsaturated fatty acids Chemical class 0.000 description 4
- 241000282414 Homo sapiens Species 0.000 description 3
- RVGRUAULSDPKGF-UHFFFAOYSA-N Poloxamer Chemical compound C1CO1.CC1CO1 RVGRUAULSDPKGF-UHFFFAOYSA-N 0.000 description 3
- 108010090804 Streptavidin Proteins 0.000 description 3
- 239000002253 acid Substances 0.000 description 3
- 150000001412 amines Chemical class 0.000 description 3
- 210000004369 blood Anatomy 0.000 description 3
- 239000008280 blood Substances 0.000 description 3
- 125000000755 henicosyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 3
- 238000003018 immunoassay Methods 0.000 description 3
- 125000002463 lignoceryl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 3
- 239000000249 polyoxyethylene sorbitan monopalmitate Substances 0.000 description 3
- 235000010483 polyoxyethylene sorbitan monopalmitate Nutrition 0.000 description 3
- 239000001818 polyoxyethylene sorbitan monostearate Substances 0.000 description 3
- 235000010989 polyoxyethylene sorbitan monostearate Nutrition 0.000 description 3
- 230000006798 recombination Effects 0.000 description 3
- 238000005215 recombination Methods 0.000 description 3
- 235000003441 saturated fatty acids Nutrition 0.000 description 3
- 210000002966 serum Anatomy 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 239000004094 surface-active agent Substances 0.000 description 3
- 125000002469 tricosyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 3
- YMXHPSHLTSZXKH-RVBZMBCESA-N (2,5-dioxopyrrolidin-1-yl) 5-[(3as,4s,6ar)-2-oxo-1,3,3a,4,6,6a-hexahydrothieno[3,4-d]imidazol-4-yl]pentanoate Chemical compound C([C@H]1[C@H]2NC(=O)N[C@H]2CS1)CCCC(=O)ON1C(=O)CCC1=O YMXHPSHLTSZXKH-RVBZMBCESA-N 0.000 description 2
- OYHQOLUKZRVURQ-NTGFUMLPSA-N (9Z,12Z)-9,10,12,13-tetratritiooctadeca-9,12-dienoic acid Chemical compound C(CCCCCCC\C(=C(/C\C(=C(/CCCCC)\[3H])\[3H])\[3H])\[3H])(=O)O OYHQOLUKZRVURQ-NTGFUMLPSA-N 0.000 description 2
- WRIDQFICGBMAFQ-UHFFFAOYSA-N (E)-8-Octadecenoic acid Natural products CCCCCCCCCC=CCCCCCCC(O)=O WRIDQFICGBMAFQ-UHFFFAOYSA-N 0.000 description 2
- QZTKDVCDBIDYMD-UHFFFAOYSA-N 2,2'-[(2-amino-2-oxoethyl)imino]diacetic acid Chemical compound NC(=O)CN(CC(O)=O)CC(O)=O QZTKDVCDBIDYMD-UHFFFAOYSA-N 0.000 description 2
- IHPYMWDTONKSCO-UHFFFAOYSA-N 2,2'-piperazine-1,4-diylbisethanesulfonic acid Chemical compound OS(=O)(=O)CCN1CCN(CCS(O)(=O)=O)CC1 IHPYMWDTONKSCO-UHFFFAOYSA-N 0.000 description 2
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 2
- LVQFQZZGTZFUNF-UHFFFAOYSA-N 2-hydroxy-3-[4-(2-hydroxy-3-sulfonatopropyl)piperazine-1,4-diium-1-yl]propane-1-sulfonate Chemical compound OS(=O)(=O)CC(O)CN1CCN(CC(O)CS(O)(=O)=O)CC1 LVQFQZZGTZFUNF-UHFFFAOYSA-N 0.000 description 2
- LQJBNNIYVWPHFW-UHFFFAOYSA-N 20:1omega9c fatty acid Natural products CCCCCCCCCCC=CCCCCCCCC(O)=O LQJBNNIYVWPHFW-UHFFFAOYSA-N 0.000 description 2
- DVLFYONBTKHTER-UHFFFAOYSA-N 3-(N-morpholino)propanesulfonic acid Chemical compound OS(=O)(=O)CCCN1CCOCC1 DVLFYONBTKHTER-UHFFFAOYSA-N 0.000 description 2
- NUFBIAUZAMHTSP-UHFFFAOYSA-N 3-(n-morpholino)-2-hydroxypropanesulfonic acid Chemical compound OS(=O)(=O)CC(O)CN1CCOCC1 NUFBIAUZAMHTSP-UHFFFAOYSA-N 0.000 description 2
- RZQXOGQSPBYUKH-UHFFFAOYSA-N 3-[[1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl]azaniumyl]-2-hydroxypropane-1-sulfonate Chemical compound OCC(CO)(CO)NCC(O)CS(O)(=O)=O RZQXOGQSPBYUKH-UHFFFAOYSA-N 0.000 description 2
- WIGIPJGWVLNDAF-UHFFFAOYSA-N 8-methyl-1-(8-methylnonoxy)nonane Chemical compound CC(C)CCCCCCCOCCCCCCCC(C)C WIGIPJGWVLNDAF-UHFFFAOYSA-N 0.000 description 2
- QSBYPNXLFMSGKH-UHFFFAOYSA-N 9-Heptadecensaeure Natural products CCCCCCCC=CCCCCCCCC(O)=O QSBYPNXLFMSGKH-UHFFFAOYSA-N 0.000 description 2
- 235000003913 Coccoloba uvifera Nutrition 0.000 description 2
- 244000060011 Cocos nucifera Species 0.000 description 2
- 235000013162 Cocos nucifera Nutrition 0.000 description 2
- PJWWRFATQTVXHA-UHFFFAOYSA-N Cyclohexylaminopropanesulfonic acid Chemical compound OS(=O)(=O)CCCNC1CCCCC1 PJWWRFATQTVXHA-UHFFFAOYSA-N 0.000 description 2
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 2
- 238000002965 ELISA Methods 0.000 description 2
- 238000008157 ELISA kit Methods 0.000 description 2
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 2
- OWXMKDGYPWMGEB-UHFFFAOYSA-N HEPPS Chemical compound OCCN1CCN(CCCS(O)(=O)=O)CC1 OWXMKDGYPWMGEB-UHFFFAOYSA-N 0.000 description 2
- GIZQLVPDAOBAFN-UHFFFAOYSA-N HEPPSO Chemical compound OCCN1CCN(CC(O)CS(O)(=O)=O)CC1 GIZQLVPDAOBAFN-UHFFFAOYSA-N 0.000 description 2
- 239000005639 Lauric acid Substances 0.000 description 2
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L Magnesium chloride Chemical compound [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 description 2
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 2
- FSVCELGFZIQNCK-UHFFFAOYSA-N N,N-bis(2-hydroxyethyl)glycine Chemical compound OCCN(CCO)CC(O)=O FSVCELGFZIQNCK-UHFFFAOYSA-N 0.000 description 2
- DBXNUXBLKRLWFA-UHFFFAOYSA-N N-(2-acetamido)-2-aminoethanesulfonic acid Chemical compound NC(=O)CNCCS(O)(=O)=O DBXNUXBLKRLWFA-UHFFFAOYSA-N 0.000 description 2
- YNLCVAQJIKOXER-UHFFFAOYSA-N N-[tris(hydroxymethyl)methyl]-3-aminopropanesulfonic acid Chemical compound OCC(CO)(CO)NCCCS(O)(=O)=O YNLCVAQJIKOXER-UHFFFAOYSA-N 0.000 description 2
- MKWKNSIESPFAQN-UHFFFAOYSA-N N-cyclohexyl-2-aminoethanesulfonic acid Chemical compound OS(=O)(=O)CCNC1CCCCC1 MKWKNSIESPFAQN-UHFFFAOYSA-N 0.000 description 2
- SEQKRHFRPICQDD-UHFFFAOYSA-N N-tris(hydroxymethyl)methylglycine Chemical compound OCC(CO)(CO)[NH2+]CC([O-])=O SEQKRHFRPICQDD-UHFFFAOYSA-N 0.000 description 2
- 239000005642 Oleic acid Substances 0.000 description 2
- ZQPPMHVWECSIRJ-UHFFFAOYSA-N Oleic acid Natural products CCCCCCCCC=CCCCCCCCC(O)=O ZQPPMHVWECSIRJ-UHFFFAOYSA-N 0.000 description 2
- 235000021314 Palmitic acid Nutrition 0.000 description 2
- WCUXLLCKKVVCTQ-UHFFFAOYSA-M Potassium chloride Chemical compound [Cl-].[K+] WCUXLLCKKVVCTQ-UHFFFAOYSA-M 0.000 description 2
- 240000008976 Pterocarpus marsupium Species 0.000 description 2
- 208000001647 Renal Insufficiency Diseases 0.000 description 2
- 229920005654 Sephadex Polymers 0.000 description 2
- 239000012507 Sephadex™ Substances 0.000 description 2
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 2
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N Sodium azide Chemical compound [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 description 2
- 235000021355 Stearic acid Nutrition 0.000 description 2
- 239000007983 Tris buffer Substances 0.000 description 2
- 239000013504 Triton X-100 Substances 0.000 description 2
- 229920004890 Triton X-100 Polymers 0.000 description 2
- 238000002835 absorbance Methods 0.000 description 2
- MBMBGCFOFBJSGT-KUBAVDMBSA-N all-cis-docosa-4,7,10,13,16,19-hexaenoic acid Chemical compound CC\C=C/C\C=C/C\C=C/C\C=C/C\C=C/C\C=C/CCC(O)=O MBMBGCFOFBJSGT-KUBAVDMBSA-N 0.000 description 2
- DTOSIQBPPRVQHS-PDBXOOCHSA-N alpha-linolenic acid Chemical compound CC\C=C/C\C=C/C\C=C/CCCCCCCC(O)=O DTOSIQBPPRVQHS-PDBXOOCHSA-N 0.000 description 2
- 235000020661 alpha-linolenic acid Nutrition 0.000 description 2
- YZXBAPSDXZZRGB-DOFZRALJSA-N arachidonic acid Chemical compound CCCCC\C=C/C\C=C/C\C=C/C\C=C/CCCC(O)=O YZXBAPSDXZZRGB-DOFZRALJSA-N 0.000 description 2
- 235000015278 beef Nutrition 0.000 description 2
- OWMVSZAMULFTJU-UHFFFAOYSA-N bis-tris Chemical compound OCCN(CCO)C(CO)(CO)CO OWMVSZAMULFTJU-UHFFFAOYSA-N 0.000 description 2
- 239000007853 buffer solution Substances 0.000 description 2
- ZCCIPPOKBCJFDN-UHFFFAOYSA-N calcium nitrate Chemical compound [Ca+2].[O-][N+]([O-])=O.[O-][N+]([O-])=O ZCCIPPOKBCJFDN-UHFFFAOYSA-N 0.000 description 2
- OSGAYBCDTDRGGQ-UHFFFAOYSA-L calcium sulfate Chemical compound [Ca+2].[O-]S([O-])(=O)=O OSGAYBCDTDRGGQ-UHFFFAOYSA-L 0.000 description 2
- 229920001577 copolymer Polymers 0.000 description 2
- 230000007423 decrease Effects 0.000 description 2
- 239000003085 diluting agent Substances 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- UKMSUNONTOPOIO-UHFFFAOYSA-N docosanoic acid Chemical compound CCCCCCCCCCCCCCCCCCCCCC(O)=O UKMSUNONTOPOIO-UHFFFAOYSA-N 0.000 description 2
- 238000004128 high performance liquid chromatography Methods 0.000 description 2
- 210000004408 hybridoma Anatomy 0.000 description 2
- 125000004029 hydroxymethyl group Chemical group [H]OC([H])([H])* 0.000 description 2
- 208000011111 hypophosphatemic rickets Diseases 0.000 description 2
- QXJSBBXBKPUZAA-UHFFFAOYSA-N isooleic acid Natural products CCCCCCCC=CCCCCCCCCC(O)=O QXJSBBXBKPUZAA-UHFFFAOYSA-N 0.000 description 2
- 201000006370 kidney failure Diseases 0.000 description 2
- 229960004488 linolenic acid Drugs 0.000 description 2
- KQQKGWQCNNTQJW-UHFFFAOYSA-N linolenic acid Natural products CC=CCCC=CCC=CCCCCCCCC(O)=O KQQKGWQCNNTQJW-UHFFFAOYSA-N 0.000 description 2
- YIXJRHPUWRPCBB-UHFFFAOYSA-N magnesium nitrate Chemical compound [Mg+2].[O-][N+]([O-])=O.[O-][N+]([O-])=O YIXJRHPUWRPCBB-UHFFFAOYSA-N 0.000 description 2
- WQEPLUUGTLDZJY-UHFFFAOYSA-N n-Pentadecanoic acid Natural products CCCCCCCCCCCCCCC(O)=O WQEPLUUGTLDZJY-UHFFFAOYSA-N 0.000 description 2
- 230000001613 neoplastic effect Effects 0.000 description 2
- QIQXTHQIDYTFRH-UHFFFAOYSA-N octadecanoic acid Chemical compound CCCCCCCCCCCCCCCCCC(O)=O QIQXTHQIDYTFRH-UHFFFAOYSA-N 0.000 description 2
- OQCDKBAXFALNLD-UHFFFAOYSA-N octadecanoic acid Natural products CCCCCCCC(C)CCCCCCCCC(O)=O OQCDKBAXFALNLD-UHFFFAOYSA-N 0.000 description 2
- WWZKQHOCKIZLMA-UHFFFAOYSA-N octanoic acid Chemical compound CCCCCCCC(O)=O WWZKQHOCKIZLMA-UHFFFAOYSA-N 0.000 description 2
- ZQPPMHVWECSIRJ-KTKRTIGZSA-N oleic acid Chemical compound CCCCCCCC\C=C/CCCCCCCC(O)=O ZQPPMHVWECSIRJ-KTKRTIGZSA-N 0.000 description 2
- 235000021313 oleic acid Nutrition 0.000 description 2
- 208000005368 osteomalacia Diseases 0.000 description 2
- 210000002381 plasma Anatomy 0.000 description 2
- 229920001451 polypropylene glycol Polymers 0.000 description 2
- 229940068977 polysorbate 20 Drugs 0.000 description 2
- FGIUAXJPYTZDNR-UHFFFAOYSA-N potassium nitrate Chemical compound [K+].[O-][N+]([O-])=O FGIUAXJPYTZDNR-UHFFFAOYSA-N 0.000 description 2
- 238000011002 quantification Methods 0.000 description 2
- 239000011541 reaction mixture Substances 0.000 description 2
- VWDWKYIASSYTQR-UHFFFAOYSA-N sodium nitrate Chemical compound [Na+].[O-][N+]([O-])=O VWDWKYIASSYTQR-UHFFFAOYSA-N 0.000 description 2
- 239000008117 stearic acid Substances 0.000 description 2
- 239000003760 tallow Substances 0.000 description 2
- TUNFSRHWOTWDNC-HKGQFRNVSA-N tetradecanoic acid Chemical compound CCCCCCCCCCCCC[14C](O)=O TUNFSRHWOTWDNC-HKGQFRNVSA-N 0.000 description 2
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- BBWMTEYXFFWPIF-CJBMEHDJSA-N (2e,4e,6e)-icosa-2,4,6-trienoic acid Chemical compound CCCCCCCCCCCCC\C=C\C=C\C=C\C(O)=O BBWMTEYXFFWPIF-CJBMEHDJSA-N 0.000 description 1
- KXVFBCSUGDNXQF-DZDBOGACSA-N (2z,4z,6z,8z,10z)-tetracosa-2,4,6,8,10-pentaenoic acid Chemical compound CCCCCCCCCCCCC\C=C/C=C\C=C/C=C\C=C/C(O)=O KXVFBCSUGDNXQF-DZDBOGACSA-N 0.000 description 1
- GYSCBCSGKXNZRH-UHFFFAOYSA-N 1-benzothiophene-2-carboxamide Chemical compound C1=CC=C2SC(C(=O)N)=CC2=C1 GYSCBCSGKXNZRH-UHFFFAOYSA-N 0.000 description 1
- FDCJDKXCCYFOCV-UHFFFAOYSA-N 1-hexadecoxyhexadecane Chemical compound CCCCCCCCCCCCCCCCOCCCCCCCCCCCCCCCC FDCJDKXCCYFOCV-UHFFFAOYSA-N 0.000 description 1
- XPALGXXLALUMLE-UHFFFAOYSA-N 2-(dimethylamino)tetradecanoic acid Chemical compound CCCCCCCCCCCCC(N(C)C)C(O)=O XPALGXXLALUMLE-UHFFFAOYSA-N 0.000 description 1
- JKXYOQDLERSFPT-UHFFFAOYSA-N 2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-(2-octadecoxyethoxy)ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethanol Chemical compound CCCCCCCCCCCCCCCCCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCO JKXYOQDLERSFPT-UHFFFAOYSA-N 0.000 description 1
- AJTVSSFTXWNIRG-UHFFFAOYSA-N 2-[bis(2-hydroxyethyl)amino]ethanesulfonic acid Chemical compound OCC[NH+](CCO)CCS([O-])(=O)=O AJTVSSFTXWNIRG-UHFFFAOYSA-N 0.000 description 1
- OGMADIBCHLQMIP-UHFFFAOYSA-N 2-aminoethanethiol;hydron;chloride Chemical compound Cl.NCCS OGMADIBCHLQMIP-UHFFFAOYSA-N 0.000 description 1
- 125000000954 2-hydroxyethyl group Chemical group [H]C([*])([H])C([H])([H])O[H] 0.000 description 1
- INEWUCPYEUEQTN-UHFFFAOYSA-N 3-(cyclohexylamino)-2-hydroxy-1-propanesulfonic acid Chemical compound OS(=O)(=O)CC(O)CNC1CCCCC1 INEWUCPYEUEQTN-UHFFFAOYSA-N 0.000 description 1
- UMCMPZBLKLEWAF-BCTGSCMUSA-N 3-[(3-cholamidopropyl)dimethylammonio]propane-1-sulfonate Chemical compound C([C@H]1C[C@H]2O)[C@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@H]([C@@H](CCC(=O)NCCC[N+](C)(C)CCCS([O-])(=O)=O)C)[C@@]2(C)[C@@H](O)C1 UMCMPZBLKLEWAF-BCTGSCMUSA-N 0.000 description 1
- XCBLFURAFHFFJF-UHFFFAOYSA-N 3-[bis(2-hydroxyethyl)azaniumyl]-2-hydroxypropane-1-sulfonate Chemical compound OCCN(CCO)CC(O)CS(O)(=O)=O XCBLFURAFHFFJF-UHFFFAOYSA-N 0.000 description 1
- 239000007991 ACES buffer Substances 0.000 description 1
- 239000004925 Acrylic resin Substances 0.000 description 1
- 229920000178 Acrylic resin Polymers 0.000 description 1
- 108700016232 Arg(2)-Sar(4)- dermorphin (1-4) Proteins 0.000 description 1
- BTBUEUYNUDRHOZ-UHFFFAOYSA-N Borate Chemical compound [O-]B([O-])[O-] BTBUEUYNUDRHOZ-UHFFFAOYSA-N 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- GHVNFZFCNZKVNT-UHFFFAOYSA-N Decanoic acid Natural products CCCCCCCCCC(O)=O GHVNFZFCNZKVNT-UHFFFAOYSA-N 0.000 description 1
- 241000283073 Equus caballus Species 0.000 description 1
- 241000588724 Escherichia coli Species 0.000 description 1
- 102000004961 Furin Human genes 0.000 description 1
- 108090001126 Furin Proteins 0.000 description 1
- 239000004471 Glycine Substances 0.000 description 1
- 239000006173 Good's buffer Substances 0.000 description 1
- 239000007995 HEPES buffer Substances 0.000 description 1
- PEEHTFAAVSWFBL-UHFFFAOYSA-N Maleimide Chemical compound O=C1NC(=O)C=C1 PEEHTFAAVSWFBL-UHFFFAOYSA-N 0.000 description 1
- 241000699666 Mus <mouse, genus> Species 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- 102000057297 Pepsin A Human genes 0.000 description 1
- 108090000284 Pepsin A Proteins 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 1
- 229920001219 Polysorbate 40 Polymers 0.000 description 1
- 239000004365 Protease Substances 0.000 description 1
- 108010076504 Protein Sorting Signals Proteins 0.000 description 1
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 description 1
- UZMAPBJVXOGOFT-UHFFFAOYSA-N Syringetin Natural products COC1=C(O)C(OC)=CC(C2=C(C(=O)C3=C(O)C=C(O)C=C3O2)O)=C1 UZMAPBJVXOGOFT-UHFFFAOYSA-N 0.000 description 1
- 239000007997 Tricine buffer Substances 0.000 description 1
- GSEJCLTVZPLZKY-UHFFFAOYSA-N Triethanolamine Chemical compound OCCN(CCO)CCO GSEJCLTVZPLZKY-UHFFFAOYSA-N 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- JAZBEHYOTPTENJ-JLNKQSITSA-N all-cis-5,8,11,14,17-icosapentaenoic acid Chemical compound CC\C=C/C\C=C/C\C=C/C\C=C/C\C=C/CCCC(O)=O JAZBEHYOTPTENJ-JLNKQSITSA-N 0.000 description 1
- OBETXYAYXDNJHR-UHFFFAOYSA-N alpha-ethylcaproic acid Natural products CCCCC(CC)C(O)=O OBETXYAYXDNJHR-UHFFFAOYSA-N 0.000 description 1
- 239000002280 amphoteric surfactant Substances 0.000 description 1
- 210000004102 animal cell Anatomy 0.000 description 1
- 239000003945 anionic surfactant Substances 0.000 description 1
- 229940114079 arachidonic acid Drugs 0.000 description 1
- 235000021342 arachidonic acid Nutrition 0.000 description 1
- CADWTSSKOVRVJC-UHFFFAOYSA-N benzyl(dimethyl)azanium;chloride Chemical compound [Cl-].C[NH+](C)CC1=CC=CC=C1 CADWTSSKOVRVJC-UHFFFAOYSA-N 0.000 description 1
- 239000007998 bicine buffer Substances 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 229960000074 biopharmaceutical Drugs 0.000 description 1
- 239000004305 biphenyl Substances 0.000 description 1
- 210000000988 bone and bone Anatomy 0.000 description 1
- 230000004097 bone metabolism Effects 0.000 description 1
- 239000006172 buffering agent Substances 0.000 description 1
- 210000004900 c-terminal fragment Anatomy 0.000 description 1
- 239000001110 calcium chloride Substances 0.000 description 1
- 229910001628 calcium chloride Inorganic materials 0.000 description 1
- 235000011148 calcium chloride Nutrition 0.000 description 1
- 235000011132 calcium sulphate Nutrition 0.000 description 1
- 239000003093 cationic surfactant Substances 0.000 description 1
- WOWHHFRSBJGXCM-UHFFFAOYSA-M cetyltrimethylammonium chloride Chemical compound [Cl-].CCCCCCCCCCCCCCCC[N+](C)(C)C WOWHHFRSBJGXCM-UHFFFAOYSA-M 0.000 description 1
- 239000012295 chemical reaction liquid Substances 0.000 description 1
- 239000003245 coal Substances 0.000 description 1
- 230000002860 competitive effect Effects 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 229960003964 deoxycholic acid Drugs 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- KCFYHBSOLOXZIF-UHFFFAOYSA-N dihydrochrysin Natural products COC1=C(O)C(OC)=CC(C2OC3=CC(O)=CC(O)=C3C(=O)C2)=C1 KCFYHBSOLOXZIF-UHFFFAOYSA-N 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 235000020669 docosahexaenoic acid Nutrition 0.000 description 1
- 229940090949 docosahexaenoic acid Drugs 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 235000020673 eicosapentaenoic acid Nutrition 0.000 description 1
- 229960005135 eicosapentaenoic acid Drugs 0.000 description 1
- JAZBEHYOTPTENJ-UHFFFAOYSA-N eicosapentaenoic acid Natural products CCC=CCC=CCC=CCC=CCC=CCCCC(O)=O JAZBEHYOTPTENJ-UHFFFAOYSA-N 0.000 description 1
- IQLUYYHUNSSHIY-HZUMYPAESA-N eicosatetraenoic acid Chemical compound CCCCCCCCCCC\C=C\C=C\C=C\C=C\C(O)=O IQLUYYHUNSSHIY-HZUMYPAESA-N 0.000 description 1
- SFNALCNOMXIBKG-UHFFFAOYSA-N ethylene glycol monododecyl ether Chemical compound CCCCCCCCCCCCOCCO SFNALCNOMXIBKG-UHFFFAOYSA-N 0.000 description 1
- 150000004665 fatty acids Chemical class 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 238000002795 fluorescence method Methods 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- BHEPBYXIRTUNPN-UHFFFAOYSA-N hydridophosphorus(.) (triplet) Chemical compound [PH] BHEPBYXIRTUNPN-UHFFFAOYSA-N 0.000 description 1
- VKOBVWXKNCXXDE-UHFFFAOYSA-N icosanoic acid Chemical compound CCCCCCCCCCCCCCCCCCCC(O)=O VKOBVWXKNCXXDE-UHFFFAOYSA-N 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 239000006166 lysate Substances 0.000 description 1
- 229910001629 magnesium chloride Inorganic materials 0.000 description 1
- 235000011147 magnesium chloride Nutrition 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
- 235000019341 magnesium sulphate Nutrition 0.000 description 1
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 1
- DVEKCXOJTLDBFE-UHFFFAOYSA-N n-dodecyl-n,n-dimethylglycinate Chemical compound CCCCCCCCCCCC[N+](C)(C)CC([O-])=O DVEKCXOJTLDBFE-UHFFFAOYSA-N 0.000 description 1
- 210000004898 n-terminal fragment Anatomy 0.000 description 1
- 229940111202 pepsin Drugs 0.000 description 1
- 239000008055 phosphate buffer solution Substances 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- 229910052698 phosphorus Inorganic materials 0.000 description 1
- 239000004417 polycarbonate Substances 0.000 description 1
- 229920000515 polycarbonate Polymers 0.000 description 1
- 229920000139 polyethylene terephthalate Polymers 0.000 description 1
- 239000005020 polyethylene terephthalate Substances 0.000 description 1
- 229920000136 polysorbate Polymers 0.000 description 1
- 229940101027 polysorbate 40 Drugs 0.000 description 1
- 229940113124 polysorbate 60 Drugs 0.000 description 1
- 229920001343 polytetrafluoroethylene Polymers 0.000 description 1
- 239000004810 polytetrafluoroethylene Substances 0.000 description 1
- 229920002635 polyurethane Polymers 0.000 description 1
- 239000004814 polyurethane Substances 0.000 description 1
- 239000004800 polyvinyl chloride Substances 0.000 description 1
- 229920000915 polyvinyl chloride Polymers 0.000 description 1
- 239000001103 potassium chloride Substances 0.000 description 1
- 235000011164 potassium chloride Nutrition 0.000 description 1
- 235000010333 potassium nitrate Nutrition 0.000 description 1
- 239000004323 potassium nitrate Substances 0.000 description 1
- OTYBMLCTZGSZBG-UHFFFAOYSA-L potassium sulfate Chemical compound [K+].[K+].[O-]S([O-])(=O)=O OTYBMLCTZGSZBG-UHFFFAOYSA-L 0.000 description 1
- 229910052939 potassium sulfate Inorganic materials 0.000 description 1
- 235000011151 potassium sulphates Nutrition 0.000 description 1
- 230000002335 preservative effect Effects 0.000 description 1
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- KCXFHTAICRTXLI-UHFFFAOYSA-N propane-1-sulfonic acid Chemical compound CCCS(O)(=O)=O KCXFHTAICRTXLI-UHFFFAOYSA-N 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 230000005855 radiation Effects 0.000 description 1
- 238000003127 radioimmunoassay Methods 0.000 description 1
- 230000009103 reabsorption Effects 0.000 description 1
- 230000000717 retained effect Effects 0.000 description 1
- 210000003296 saliva Anatomy 0.000 description 1
- FHHPUSMSKHSNKW-SMOYURAASA-M sodium deoxycholate Chemical compound [Na+].C([C@H]1CC2)[C@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@H]([C@@H](CCC([O-])=O)C)[C@@]2(C)[C@@H](O)C1 FHHPUSMSKHSNKW-SMOYURAASA-M 0.000 description 1
- 239000004402 sodium ethyl p-hydroxybenzoate Substances 0.000 description 1
- 235000010344 sodium nitrate Nutrition 0.000 description 1
- 239000004317 sodium nitrate Substances 0.000 description 1
- 229910052938 sodium sulfate Inorganic materials 0.000 description 1
- 235000011152 sodium sulphate Nutrition 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 210000005239 tubule Anatomy 0.000 description 1
- 210000002700 urine Anatomy 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/531—Production of immunochemical test materials
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/475—Growth factors; Growth regulators
- C07K14/50—Fibroblast growth factor [FGF]
Definitions
- the present invention relates to a stabilizer for fibroblast growth factor-23 (hereinafter referred to as FGF-23), a method for stabilizing FGF-23, a method for storing FGF-23, and a reagent for measuring FGF-23.
- FGF-23 fibroblast growth factor-23
- FGF-23 is a member of the fibroblast growth factor (FGF) family, is produced mainly in bone tissue, acts on the kidney, and inhibits reabsorption of phosphorus in the renal tubule.
- FGF fibroblast growth factor
- FGF-23 is a protein consisting of 251 amino acids, of which 24 amino acids at the N-terminus are signal peptides, and secreted FGF-23 is considered to consist of 227 amino acids.
- Some FGF-23s are processed between the 179th amino acid Arg and the 180th amino acid Ser by furin or the like, which is a kind of protease (see Non-Patent Documents 2 and 3).
- the full length FGF-23 consisting of 227 amino acids, which is not processed, has biological activities such as a decrease in blood phosphorous concentration, whereas the N-terminal and C-terminal fragments after this processing are It is reported to be inactive (see Non-Patent Documents 2 and 3).
- FGF-23 is very unstable, and when an aqueous medium containing FGF-23 is stored in a container, the concentration of FGF-23 in the aqueous medium gradually decreases. Therefore, there are problems that it is difficult to prepare an accurate calibration curve and that it is difficult to accurately measure FGF-23 in a sample based on the calibration curve.
- An object of the present invention is to provide an FGF-23 stabilizer, an FGF-23 stabilization method, an FGF-23 storage method, and an FGF-23 measurement, which enable accurate measurement of FGF-23 in a sample. It is to provide a reagent for use.
- the inventor of the present invention diligently studied to solve the above problems, and found the finding that FGF-23 is stabilized by a polyoxyethylene-based nonionic surfactant, and completed the present invention. . That is, the present invention relates to the following [1] to [17].
- a stabilizer for FGF-23 which contains a polyoxyethylene-based nonionic surfactant as an active ingredient.
- the polyoxyethylene nonionic surfactant is selected from the group consisting of polyoxyethylene sorbitan fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene alkylphenyl ether, and polyoxyethylene alkylamine.
- the stabilizer according to [1] above which is at least one nonionic surfactant.
- the stabilizer according to [1] or [2] further containing albumin.
- the stabilizer according to [3], wherein the albumin is bovine serum albumin.
- a method for stabilizing FGF-23 wherein a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
- the polyoxyethylene nonionic surfactant is selected from the group consisting of polyoxyethylene sorbitan fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene alkylphenyl ether, and polyoxyethylene alkylamine.
- the stabilization method according to [5] wherein the stabilization method is at least one nonionic surfactant.
- the plastic container is a container selected from the group consisting of a polyethylene container, a polypropylene container, and a polystyrene container.
- a method for storing FGF-23 wherein a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
- the polyoxyethylene-based nonionic surfactant is selected from the group consisting of polyoxyethylene sorbitan fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene alkylphenyl ether, and polyoxyethylene alkylamine.
- the storage method according to [11] above which is at least one nonionic surfactant.
- a reagent for measuring FGF-23 comprising the stabilizer for FGF-23 according to any one of [1] to [4].
- an FGF-23 stabilizer an FGF-23 stabilization method, an FGF-23 storage method, and an FGF-23 measurement reagent that enable accurate measurement of FGF-23 in a sample Is provided.
- the stabilizer for FGF-23 of the present invention contains a polyoxyethylene nonionic surfactant as an active ingredient. Further, the FGF-23 stabilizer of the present invention may contain albumin in addition to the polyoxyethylene nonionic surfactant.
- the FGF-23 stabilizer of the present invention has an effect of stably holding FGF-23 in an aqueous FGF-23 solution contained in a container.
- stable FGF-23 means that the FGF-23 concentration in the FGF-23 aqueous solution contained in the container is substantially constant while the FGF-23 aqueous solution is stored in the container. It means that it is maintained.
- the method for measuring the FGF-23 concentration in the FGF-23 aqueous solution contained in the container may be any method as long as it can accurately measure FGF-23.
- a well-known FGF-based immunoassay method may be used. 23 measurement methods and the like. Examples of the immunological measurement method include immunological measurement methods described later.
- FGF-23 stabilized by the stabilizer of the present invention may be produced by a gene recombination method or collected from a biological sample.
- Examples of the gene recombination method used for producing FGF-23 by a gene recombination method include a method of producing FGF-23 in a transformant obtained using Escherichia coli as an host and animal cells. .
- As a method for collecting FGF-23 from a biological sample for example, a method for separating and purifying FGF-23 from a biological sample such as blood, plasma, or serum that may contain FGF-23.
- FGF-23 can also be produced using a known method (for example, see WO2012 / 029837).
- the FGF-23 aqueous solution is a solution in which FGF-23 is dissolved in an aqueous medium.
- the aqueous medium is not particularly limited as long as it is capable of stably holding FGF-23, and examples thereof include deionized water, distilled water, and a buffer solution. Among these, a buffer solution is preferable.
- buffering agents include, for example, 2-morpholinoethanesulfonic acid (MES), bis (2-hydroxyethyl) iminotris (hydroxymethyl) methane (Bis-Tris), N- (2-acetamido) iminodiacetic acid (ADA) Piperazine-N, N'-bis (2-ethanesulfonic acid) (PIPES), N- (2-acetamido) -2-aminoethanesulfonic acid (ACES), 3-morpholino-2-hydroxypropanesulfonic acid (MOPSO) ), N, N-bis (2-hydroxyethyl) -2-aminoethanesulfonic acid (BES), 3-morpholinopropanesulfonic acid (MOPS), N- [tris (hydroxymethyl) methyl] -2-aminoethanesulf
- MES 2-morpholinoethanesulfonic acid
- Bis-Tris bis (2-hydroxyethyl) iminotris (hydroxy
- the container in the present invention is a container for storing an FGF-23 aqueous solution, and is a container used for storing the FGF-23 aqueous solution.
- Examples of the container in the present invention include a plastic container.
- the plastic container means a container whose surface that comes into contact with the FGF-23-containing sample is made of plastic, and is not only a container whose whole container is made of plastic, but is a container whose only surface that comes into contact with the FGF-23-containing sample is made of plastic. are also included.
- plastic container examples include a polyethylene container, a polypropylene container, a polystyrene container, a polyvinyl chloride container, a polyurethane container, a polytetrafluoroethylene container, a polyethylene terephthalate container, an acrylic resin container, and a polycarbonate container.
- a polyethylene container, a polypropylene container, and a polystyrene container are preferable.
- the polyoxyethylene nonionic surfactant in the present invention is not particularly limited as long as it is a nonionic surfactant having a polyoxyethylene structure.
- polyoxyethylene sorbitan fatty acid ester hereinafter referred to as POE sorbitan fatty acid ester.
- Polyoxyethylene alkyl ether (hereinafter referred to as POE alkyl ether), polyoxyethylene alkyl phenyl ether (hereinafter referred to as POE alkyl phenyl ether), polyoxyethylene alkyl amine (hereinafter referred to as POE alkyl amine), polyoxyethylene Polyoxypropylene copolymer (hereinafter referred to as POE / POP copolymer), polyoxyethylene / polyoxypropylene alkyl ether (hereinafter referred to as POE / POP alkyl ether), polyoxyethylene Ethylene-polyoxypropylene alkylphenyl ether (hereinafter, POE-called POP alkyl phenyl ether), and the like.
- POE alkyl ether Polyoxyethylene alkyl ether
- POE alkyl phenyl ether polyoxyethylene alkyl phenyl ether
- POE alkyl amine polyoxyethylene alkyl amine
- POE sorbitan fatty acid esters POE alkyl ethers, POE alkylphenyl ethers, POE alkyl amines are preferred.
- the polyoxyethylene nonionic surfactant can be used alone or in combination of two or more.
- commercially available products can also be used as the polyoxyethylene-based nonionic surfactant.
- Examples of the fatty acid in the POE sorbitan fatty acid ester include saturated or unsaturated fatty acids having 8 to 24 carbon atoms, and among these, saturated or unsaturated fatty acids having 12 to 18 carbon atoms are preferable.
- Examples of saturated or unsaturated fatty acids having 8 to 24 carbon atoms include octanoic acid, decanoic acid, lauric acid, myristic acid, palmitic acid, stearic acid, oleic acid, linoleic acid, linolenic acid, eicosatrienoic acid, arachidonic acid, Examples include icosanoic acid, eicosatetraenoic acid, eicosapentaenoic acid, docosanic acid, docosahexaenoic acid, tetradocosanoic acid, tetracosapentaenoic acid, and the like.
- saturated or unsaturated fatty acid having 12 to 18 carbon atoms include lauric acid, myristic acid, palmitic acid, stearic acid, oleic acid, linoleic acid, and linolenic acid.
- POE sorbitan fatty acid ester include polyoxyethylene sorbitan monolaurate, polyoxyethylene sorbitan monooleate, polyoxyethylene sorbitan monostearate, polyoxyethylene sorbitan monopalmitate and the like.
- POE sorbitan fatty acid esters examples include polysorbate 80 (Tween 80) (polyoxyethylene sorbitan monooleate), polysorbate 40 (Tween 40) (polyoxyethylene sorbitan monopalmitate), polysorbate 60 (Tween 60) ( Polyoxyethylene sorbitan monostearate), polysorbate 20 (Tween 20) (polyoxyethylene sorbitan monolaurate) (above, Sigma-Aldrich), BLAUNON OT-21 (polyoxyethylene sorbitan monooleate) (Aoki oil) Manufactured by Kogyo Co., Ltd.).
- alkyl in the POE alkyl ether examples include alkyl having 8 to 24 carbon atoms, and among these, alkyl having 10 to 20 carbon atoms is preferable.
- alkyl having 8 to 24 carbon atoms include octyl, isooctyl, nonyl, decyl, isodecyl, undecyl, dodecyl (lauryl), tridecyl, tetradecyl (myristyl), pentadecyl, hexadecyl (cetyl), heptadecyl, octadecyl (stearyl), oleyl , Nonadecyl, icosyl, heneicosyl, docosyl (behenyl), tricosyl, tetracosyl and the like.
- alkyl having 10 to 20 carbon atoms examples include decyl, isodecyl, undecyl, dodecyl (lauryl), tridecyl, tetradecyl (myristyl), pentadecyl, hexadecyl (cetyl), heptadecyl, octadecyl (stearyl), oleyl, nonadecyl, icosyl and the like. Can be mentioned.
- POE alkyl ethers include, for example, Brij35 (polyoxyethylene lauryl ether), Brij99 (polyoxyethylene oleyl ether), Brij56 (polyoxyethylene cetyl ether), Brij78 (polyoxyethylene stearyl ether) (above, Sigma- Aldrich), Nikkor BC-23 (polyoxyethylene cetyl ether) (Nikko Chemicals), nonion P-210, nonion P-213 (above, polyoxyethylene cetyl ether), nonion K-220 (polyoxyethylene) Lauryl ether), nonion E-205, nonion E-215, nonion E-230 (more, polyoxyethylene oleyl ether), nonion S-215, nonion S-220 (more, polyoxyethylene stearyl ether) (more, day Oil company), Emulgen 120 (polyoxyethylene lauryl ether), Margen 220 (polyoxyethylene cetyl ether), Emulgen 320P (polyoxyethylene
- Examples of the alkyl in the POE alkyl phenyl ether include alkyl having 8 to 9 carbon atoms. Examples of the alkyl having 8 to 9 carbon atoms include octyl and nonyl. Examples of commercially available POE alkylphenyl ethers include Triton X-100 (manufactured by Sigma-Aldrich) (polyoxyethylene octylphenyl ether), Emulgen 810 (polyoxyethylene octylphenyl ether), Emulgen 910, Emulgen 930 (above, Polyoxyethylene nonylphenyl ether) (above, manufactured by Kao Corporation), Nikkor OP-10 (polyoxyethylene octylphenyl ether), Nikkor NP-10 (polyoxyethylene nonylphenyl ether) (above, manufactured by Nikko Chemicals), Nonion HS-210, Nonion HS-215, Nonion HS-220 (above, poly
- alkyl in the POE alkylamine examples include alkyl having 8 to 24 carbon atoms, and among these, alkyl having 10 to 20 carbon atoms is preferable.
- alkyl having 8 to 24 carbon atoms include octyl, isooctyl, nonyl, decyl, isodecyl, undecyl, dodecyl (lauryl), tridecyl, tetradecyl (myristyl), pentadecyl, hexadecyl (cetyl), heptadecyl, octadecyl (stearyl), oleyl , Nonadecyl, icosyl, heneicosyl, docosyl (behenyl), tricosyl, tetracosyl and the like.
- alkyl having 10 to 20 carbon atoms examples include decyl, isodecyl, undecyl, dodecyl (lauryl), tridecyl, tetradecyl (myristyl), pentadecyl, hexadecyl (cetyl), heptadecyl, octadecyl (stearyl), oleyl, nonadecyl, icosyl and the like. Can be mentioned.
- POE alkylamines include, for example, Neimine L-207 (polyoxyethylene laurylamine), Neimine S-210, Naimine S-215, Naimine S-220 (above, polyoxyethylene stearylamine), Naimine T2-210 , Naimine T2-230 [above, polyoxyethylene alkyl (beef tallow) amine], Naimine F-215 [polyoxyethylene alkyl (coconut) amine] (above, NOF Corporation), BLAUNON L-210, BLAUNON L- 230 (above, polyoxyethylene laurylamine), BLAUNON S-210, BLAUNON S-215, BLAUNON S-220, BLAUNON S-230 (above, polyoxyethylene stearylamine), BLAUNON S-210T, BLAUNON S-215T, BLAUNON S-220T, BLAUNON S-230T [above, polyoxyethylene alkyl (beef tallow) amine]
- Examples of commercially available POE / POP copolymers include Pronon 201, Pronon 204, and Pronon 208 (above, NOF Corporation).
- alkyl in the POE / POP alkyl ether examples include alkyl having 8 to 24 carbon atoms, and among these, alkyl having 10 to 20 carbon atoms is preferable.
- alkyl having 8 to 24 carbon atoms include octyl, isooctyl, nonyl, decyl, isodecyl, undecyl, dodecyl (lauryl), tridecyl, tetradecyl (myristyl), pentadecyl, hexadecyl (cetyl), heptadecyl, octadecyl (stearyl), oleyl , Nonadecyl, icosyl, heneicosyl, docosyl (behenyl), tricosyl, tetracosyl and the like.
- alkyl having 10 to 20 carbon atoms examples include decyl, isodecyl, undecyl, dodecyl (lauryl), tridecyl, tetradecyl (myristyl), pentadecyl, hexadecyl (cetyl), heptadecyl, octadecyl (stearyl), oleyl, nonadecyl, icosyl and the like. Can be mentioned.
- Examples of commercially available POE / POP alkyl ethers include Fine Surf IDEP-802, Wanda Surf ID-90 (above, polyoxyethylene polyoxypropylene isodecyl ether), Wanda Surf RL-80, Wanda Surf RL-100, Wanda Surf RL-140 (above, polyoxyethylene polyoxypropylene lauryl ether) (above, manufactured by Aoki Yushi Kogyo Co., Ltd.) and the like.
- Examples of commercially available POE / POP alkylphenyl ethers include Emulgen L40 (polyoxyethylene polyoxypropylene octylphenyl ether) (manufactured by Kao Corporation).
- the HLB value (Hydrophile-Lipophile Balance value) of the polyoxyethylene-based nonionic surfactant in the present invention is not particularly limited as long as it is an HLB value capable of stabilizing FGF-23, and is usually 12 to 20, 12.5 ⁇ 19 are preferred, and 13-18 are more preferred.
- the concentration of the polyoxyethylene-based nonionic surfactant to be used is not particularly limited as long as it is a concentration capable of stabilizing FGF-23, and is usually 0.00001 to 10 w / v%, Is preferred.
- albumin in the present invention examples include bovine, horse, sheep, human, mouse, and rabbit-derived albumin, and among these, bovine serum albumin (BSA) is preferable. Also, albumin produced by genetic engineering techniques can be used. In the present invention, two or more types of albumin can be used in combination.
- the concentration of albumin used is not particularly limited as long as it can stabilize FGF-23, and is usually 0.01 to 5 w / v%, preferably 0.05 to 1 w / v%.
- the stabilizer of the present invention may contain a chelating agent, salts, preservatives and the like.
- the chelating agent include ethylenediaminetetraacetic acid (EDTA).
- the salts include sodium chloride, potassium chloride, sodium sulfate, potassium sulfate, sodium nitrate, potassium nitrate, magnesium chloride, calcium chloride, magnesium sulfate, calcium sulfate, magnesium nitrate, and calcium nitrate.
- the preservative include sodium azide and bioacet.
- the stability of FGF-23 by the FGF-23 stabilizer of the present invention can be evaluated by the following method.
- a solution X containing FGF-23 is prepared in a container.
- FGF-23 concentration in the solution X immediately after preparation [CX (immediately after preparation) is determined.
- the solution X is stored at 4 ° C. for 7 days, and the FGF-23 concentration [CX (after storage) ] in the solution X after the storage is determined.
- the residual ratio (%) of FGF-23 to the solution X is calculated from the following formula (I).
- the concentration of FGF-23 can be determined by any method as long as it can accurately measure FGF-23.
- the FGF-23 concentration is determined by using a known FGF-23 measurement method by immunoassay. be able to.
- the immunological measurement method include a sandwich method and a competitive method based on the measurement principle, and examples include an absorbance method, a chemiluminescence method, a fluorescence method, and a radioimmunoassay method based on the detection method. . Therefore, the FGF-23 concentration can be determined using the absorbance, emission intensity, fluorescence intensity, radiation intensity, etc. corresponding to the FGF-23 concentration.
- FGF-23 measurement methods by immunoassay include a method using a commercially available FGF-23 measurement kit.
- Examples of commercially available FGF-23 measurement kits include Human Intact FGF-23 ELISA KIT (Immutopics), Human FGF-23 ELISA KIT (Merck Millipore), and FGF-23 measurement reagent (Kinos).
- the method for stabilizing FGF-23 of the present invention is characterized in that a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant. It is. Specific embodiments of the stabilization method will be described below.
- ⁇ Stabilization method 1 A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a container containing a sample containing FGF-23.
- ⁇ Stabilization method 2 A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a sample containing FGF-23 to a container containing a polyoxyethylene-based nonionic surfactant.
- ⁇ Stabilization method 3 A method for stabilizing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant to a container containing an aqueous medium containing a sample containing FGF-23.
- ⁇ Stabilization method 4 A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
- the method for stabilizing FGF-23 is characterized in that a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene nonionic surfactant and albumin. Is the method. Specific embodiments of the stabilization method will be described below.
- ⁇ Stabilization method 5 A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene nonionic surfactant and albumin to a container containing a sample containing FGF-23.
- ⁇ Stabilization method 6 A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing albumin to a container containing FGF-23 and a container containing a polyoxyethylene-based nonionic surfactant.
- ⁇ Stabilization method 7 A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a sample containing FGF-23 and a container containing albumin.
- ⁇ Stabilization method 8 A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 and an aqueous medium containing albumin to a container containing a polyoxyethylene-based nonionic surfactant.
- ⁇ Stabilization method 9 A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a sample containing FGF-23 to a container containing a polyoxyethylene-based nonionic surfactant and albumin.
- ⁇ Stabilization method 10 A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 and an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a container containing albumin.
- ⁇ Stabilization method 11 A method for stabilizing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant and albumin to a container containing an aqueous medium containing a sample containing FGF-23.
- ⁇ Stabilization method 12 A method for stabilizing FGF-23, comprising a step of adding albumin to a container containing a sample containing FGF-23 and an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
- ⁇ Stabilization method 13 A method for stabilizing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant to a container containing a sample containing FGF-23 and an aqueous medium containing albumin.
- ⁇ Stabilization method 14 A method for stabilizing FGF-23, comprising the step of adding a sample containing FGF-23 and albumin to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
- ⁇ Stabilization method 15 A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 to a container containing an aqueous medium containing polyoxyethylene-based nonionic surfactant and albumin.
- ⁇ Stabilization method 16 A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 and a polyoxyethylene-based nonionic surfactant to a container containing an aqueous medium containing albumin.
- the sample containing FGF-23 is not particularly limited as long as it is a sample containing FGF-23, and examples thereof include biological samples containing FGF-23 and FGF-23 standard solutions.
- the biological sample examples include blood, serum, plasma, urine, saliva, spinal fluid, stool, and the like.
- the FGF-23 standard solution is an aqueous solution containing a known concentration of FGF-23, and is used for preparing a calibration curve necessary for quantifying FGF-23 in a sample obtained from a living body.
- Examples of FGF-23 used in the FGF-23 standard solution include the aforementioned FGF-23.
- Containers used in the method for stabilizing FGF-23 of the present invention, polyoxyethylene-based nonionic surfactants, and albumin include, for example, the aforementioned containers, polyoxyethylene-based nonionic surfactants And albumin.
- the content of FGF-23 in the biological sample is usually 5 to 10,000 ⁇ pg / mL, and the calibration curve is usually prepared in the FGF-23 standard solution in this concentration range. It is done using.
- the polyoxyethylene-based nonionic surfactant and its concentration, and albumin and its concentration used in the method for stabilizing FGF-23 of the present invention include the aforementioned polyoxyethylene-based nonionic surfactants. Surfactant and its concentration, and albumin and its concentration are mentioned, respectively.
- the stabilization period in the FGF-23 stabilization method of the present invention is not particularly limited as long as FGF-23 is stably held, and is usually 7 days to 1 year. Further, in the method for stabilizing FGF-23 of the present invention, a sample containing FGF-23 is placed in a container in a polyoxyethylene-based nonionic surfactant or a polyoxyethylene-based nonionic interface.
- the temperature for coexisting with the active agent and albumin is not particularly limited as long as FGF-23 is stably maintained, and is usually -5 to 45 ° C, preferably 0 to 30 ° C, and preferably 2 to 10 ° C. Particularly preferred.
- chelating agents, salts, preservatives and the like may be added to the FGF-23 aqueous solution.
- chelating agents, salts, and preservatives include the aforementioned chelating agents, salts, and preservatives.
- the method for preserving FGF-23 of the present invention is a method characterized in that a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant. is there. Specific embodiments of the storage method will be described below.
- Storage method 1 A method for storing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a container containing a sample containing FGF-23.
- ⁇ Storage method 2 A method for storing FGF-23, comprising a step of adding an aqueous medium containing a sample containing FGF-23 to a container containing a polyoxyethylene-based nonionic surfactant.
- Storage method 3 A method for storing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant to a container containing an aqueous medium containing a sample containing FGF-23.
- Save method 4 A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
- a method for storing FGF-23 is characterized in that a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant and albumin. It is. Specific embodiments of the storage method will be described below.
- ⁇ Storage method 6 A method for storing FGF-23, comprising a step of adding an aqueous medium containing albumin to a container containing FGF-23 and a container containing a polyoxyethylene-based nonionic surfactant.
- Storage method 7 A method for storing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a sample containing FGF-23 and a container containing albumin.
- ⁇ Storage method 8 A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 and an aqueous medium containing albumin to a container containing a polyoxyethylene-based nonionic surfactant.
- ⁇ Storage method 9 A method for storing FGF-23, comprising a step of adding an aqueous medium containing a sample containing FGF-23 to a container containing a polyoxyethylene-based nonionic surfactant and albumin.
- Storage method 10 A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 and an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a container containing albumin.
- ⁇ Storage method 11 A method for storing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant and albumin to a container containing an aqueous medium containing a sample containing FGF-23.
- ⁇ Storage method 12 A method for storing FGF-23, comprising a step of adding albumin to a container containing a sample containing FGF-23 and an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
- ⁇ Storage method 13 A method for storing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant to a container containing a sample containing FGF-23 and an aqueous medium containing albumin.
- ⁇ Storage method 14 A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 and albumin to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
- ⁇ Storage method 15 A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant and albumin.
- ⁇ Storage method 16 A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 and a polyoxyethylene-based nonionic surfactant to a container containing an aqueous medium containing albumin.
- FGF-23 and its concentration, polyoxyethylene nonionic surfactant and its concentration, and albumin and its concentration used in the method for storing FGF-23 of the present invention include the above-mentioned FGF.
- FGF-23 and its concentration in a stabilizer of ⁇ 23 examples thereof include FGF-23 and its concentration in a stabilizer of ⁇ 23, polyoxyethylene nonionic surfactant and its concentration, and albumin and its concentration.
- the storage period in the FGF-23 storage method of the present invention is not particularly limited as long as FGF-23 is stably stored, and is usually 7 days to 1 year.
- the storage temperature in the method for storing FGF-23 of the present invention is not particularly limited as long as FGF-23 is stably stored, and is usually -5 to 45 ° C, preferably 0 to 30 ° C. 2 to 10 ° C. is particularly preferable.
- chelating agents, salts, preservatives, and the like may be added to the FGF-23 aqueous solution.
- chelating agents, salts, and preservatives include the aforementioned chelating agents, salts, and preservatives.
- the reagent for measuring FGF-23 of the present invention contains the stabilizer for FGF-23 of the present invention.
- the reagent for measuring FGF-23 of the present invention may contain other components other than the stabilizer for FGF-23 of the present invention.
- the stabilizer for FGF-23 of the present invention in the reagent for measuring FGF-23 of the present invention, is placed in a container independent of other components and independent of the container in which other components are placed. However, it may be contained in another FGF-23 measurement reagent and placed in the same container.
- the FGF-23 stabilizer of the present invention is preferably placed in the same container as the other components.
- the “other components” include a sample diluent, a magnetic particle suspension, a biotin-conjugated anti-FGF-23 antibody-containing reagent, an alkaline phosphatase-labeled anti-FGF-23 antibody fragment-containing reagent, and a standard solution for FGF-23 measurement.
- a form in which the FGF-23 stabilizer of the present invention is contained in a sample diluent and / or a standard solution for FGF-23 measurement is preferable.
- FGF-23 in the standard product is stabilized, so that accurate calibration is possible.
- a line can be created, and an accurate measurement of FGF-23 based on an accurate calibration curve is possible.
- Polysorbate 80 polyoxyethylene sorbitan monooleate; HLB 15.0) (Sigma-Aldrich), Polysorbate 20 (polyoxyethylene sorbitan monolaurate; HLB 16.7) (Sigma-Aldrich), Brij 35 (polyoxyethylene) Lauryl ether; HLB 16.9) (manufactured by Sigma-Aldrich), Brij 99 (polyoxyethylene oleyl ether; HLB 15.3) (manufactured by Sigma-Aldrich), Triton X-100 (polyoxyethylene octylphenyl ether; HLB 16.9) ( Sigma-Aldrich), Naimine S-220 (polyoxyethylene octadecylamine; HLB 15.4) (manufactured by NOF Corporation), Nissan Cation PB-300 (hexadecyltrimethylammonium chloride) (manufactured by NOF Corporation), Nissan Cation F2 -35R [alkyl (coconut) dimethyl
- FGF-23 solution (solutions A1 to A6; solutions a1 to a6)
- FGF-23 solution (solutions A1 to A6) having the following composition A6; Solutions a1 to a6) were prepared.
- Phosphate buffer 10 mmol / L (pH7.4)
- FGF-23 1 ⁇ g / mL Sodium chloride 140 mmol / L Surfactant (See Table 1) 0.0005%
- FGF-23 solution (solution a0) having the following composition was prepared.
- FGF-23 1 ⁇ g / mL Sodium chloride 140 mmol / L
- the solution a0 has the same composition as the solutions A1 to A6 and the solutions a1 to a6 except that the surfactant is not included.
- antigen dilution liquid having the following composition was prepared.
- Phosphate buffer solution 20 mmol / L (pH6.5)
- Sodium chloride 300 mmol / L BSA 0.2 (w / v)%
- FGF-23 quantification kit FGF-23 quantification comprising the following magnetic particle suspension, biotin-conjugated anti-FGF-23 antibody-containing reagent, and alkaline phosphatase-labeled anti-FGF-23 antibody fragment-containing reagent. A kit was prepared.
- Magnetic particles As magnetic particles, commercially available magnetic particles (Dynabeads MyOne Streptavidin; manufactured by Dynal) bound with streptavidin were used to prepare a magnetic particle suspension having the following composition.
- MES pH 6.5
- 50 mmol / L Streptavidin-coupled magnetic particles 0.75 g / L BSA 0.1% Sodium chloride 0.1 mol / L
- biotin-conjugated anti-FGF-23 monoclonal antibody a biotin-conjugated anti-FGF-23 antibody-containing reagent having the following composition was prepared.
- MES pH 6.5
- Anti-FGF-23 monoclonal antibody 5 ⁇ g / mL BSA 0.1% Sodium chloride 0.1 mol / L
- the obtained F (ab ′) 2 was reduced with 2-mercaptoethylamine hydrochloride (manufactured by Nacalai Tesque), and then a HPLC system using a G3000SW column (manufactured by Tosoh Corporation; aperture: 21.5 mm; length: 60 cm) Fab 'was separated by (manufactured by Hitachi, Ltd.).
- the obtained Fab ′ and alkaline phosphatase were bound by the maleimide method according to the following procedure.
- maleimide-forming reagent Sulfo-HMCS manufactured by Dojindo Laboratories
- alkaline phosphatase was maleimidated
- the reaction mixture was subjected to Sephadex G-25 column (GE Health Science Japan) to make unreacted Sulfo- HMCS was removed to obtain maleimidated alkaline phosphatase.
- alkaline phosphatase labeled Fab ′ antibody The prepared maleimidated alkaline phosphatase and Fab ′ were mixed to prepare an alkaline phosphatase labeled Fab ′ antibody.
- an alkaline phosphatase labeled anti-FGF-23 antibody fragment solution having the following composition was prepared.
- MES pH 6.5
- Alkaline phosphatase-labeled anti-FGF-23 antibody fragment 7 ⁇ g / mL BSA 0.1% Sodium chloride 0.1 mol / L
- Antigen dilution solution of each of the following concentrations of FGF-23 was prepared as a measurement sample: 10,000 pg / mL; 3,000 pg / mL; 1,000 pg / mL; 300 pg / mL; 100 pg / mL; 50 pg / mL; 5 pg / mL.
- a 20 mmol / L phosphate buffer (pH 6.5) containing 0.2 (w / v)% BSA and 300 mmol / L sodium chloride was used as the antigen dilution.
- the prepared measurement sample (10 ⁇ L) was added to the reaction cuvette, and then the magnetic particle suspension prepared in (4) above, the biotin-conjugated anti-FGF-23 antibody solution, and the alkaline phosphatase-labeled anti-FGF-23 antibody fragment 30 ⁇ L of each constituent reagent of the solution was added and stirred, and reacted at 37 ° C. for 10 minutes.
- the magnetic particles in the reaction mixture are collected by magnetic force to remove the reaction liquid other than the magnetic particles, and at the same time, the washing liquid [10% mmol / L phosphate buffer (pH 7.7 containing 0.05% Tween20 and 140% mmol / L sodium chloride). 4)], the magnetic particles were washed five times.
- a luminescent substrate solution (100 ⁇ L) mainly composed of 9- (4-chlorophenylthiophosphoryloxymethylidene) -10-methylacridan disodium salt was added and stirred, and the luminescence intensity of the generated light was measured. .
- FGF-23 solution (solution A7) having the following composition was prepared.
- Phosphate buffer 10 mmol / L (pH7.4)
- FGF-23 1 ⁇ g / mL Sodium chloride 140 mmol / L
- FGF-23 solution (solution B) having the following composition was prepared.
- Phosphate buffer 10 mmol / L (pH7.4)
- FGF-23 1 ⁇ g / mL Sodium chloride 140 mmol / L
- an FGF-23 stabilizer effective for diagnosis of diseases such as hypophosphatemic rickets, neoplastic osteomalacia and renal failure, a method for stabilizing FGF-23, and a method for storing FGF-23 And a reagent for measuring FGF-23.
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Molecular Biology (AREA)
- Chemical & Material Sciences (AREA)
- Biomedical Technology (AREA)
- Urology & Nephrology (AREA)
- Hematology (AREA)
- Cell Biology (AREA)
- Analytical Chemistry (AREA)
- Biotechnology (AREA)
- Pathology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Peptides Or Proteins (AREA)
- Medicinal Preparation (AREA)
Abstract
The present invention provides an agent for stabilizing fibroblast growth factor 23 (referred to below as "FGF23"), a method for stabilizing FGF23, a method for retaining FGF23, and a reagent for measuring FGF23. This agent for stabilizing FGF23 contains a polyoxyethylene-based non-ionic surfactant as an active ingredient. This method for stabilizing FGF23 and method for retaining FGF23 are characterized in that a specimen containing FGF23 is made to coexist in a container with an aqueous medium that contains a polyoxyethylene-based non-ionic surfactant. This reagent for measuring FGF23 contains this agent for stabilizing FGF23.
Description
本発明は、線維芽細胞増殖因子－２３（以下、ＦＧＦ－２３と記す）の安定化剤、ＦＧＦ－２３の安定化方法、ＦＧＦ－２３の保存方法、及び、ＦＧＦ－２３測定用試薬に関する。
The present invention relates to a stabilizer for fibroblast growth factor-23 (hereinafter referred to as FGF-23), a method for stabilizing FGF-23, a method for storing FGF-23, and a reagent for measuring FGF-23.
ＦＧＦ－２３は、線維芽細胞増殖因子（ＦＧＦ）ファミリーの一員であり、主として骨組織で産生され、腎臓に作用し、腎尿細管でのリンの再吸収を阻害する。近年、低リン血症性くる病、腫瘍性骨軟化症、腎不全等の疾患へのＦＧＦ－２３の関与が示俊されている（特許文献１；非特許文献１参照）。
FGF-23 is a member of the fibroblast growth factor (FGF) family, is produced mainly in bone tissue, acts on the kidney, and inhibits reabsorption of phosphorus in the renal tubule. In recent years, the involvement of FGF-23 in diseases such as hypophosphatemic rickets, neoplastic osteomalacia, and renal failure has been demonstrated (see Patent Document 1; Non-Patent Document 1).
ＦＧＦ－２３は、251個のアミノ酸からなる蛋白であり、このうちＮ端24個のアミノ酸はシグナルペプチドであり、分泌されるＦＧＦ－２３は227個のアミノ酸からなるものと考えられている。一部のＦＧＦ－２３は、プロテアーゼの１種であるfurin等により179番目のアミノ酸Argと180番目のアミノ酸Serとの間でプロセッシングを受ける（非特許文献２、３参照）。プロセッシングを受けない、227個のアミノ酸からなる全長ＦＧＦ－２３は、血中リン濃度の低下等の生物活性を有するのに対して、このプロセッシングを受けた後のＮ端、及び、Ｃ端フラグメントは不活性であることが報告されている（非特許文献２、３参照）。
FGF-23 is a protein consisting of 251 amino acids, of which 24 amino acids at the N-terminus are signal peptides, and secreted FGF-23 is considered to consist of 227 amino acids. Some FGF-23s are processed between the 179th amino acid Arg and the 180th amino acid Ser by furin or the like, which is a kind of protease (see Non-Patent Documents 2 and 3). The full length FGF-23 consisting of 227 amino acids, which is not processed, has biological activities such as a decrease in blood phosphorous concentration, whereas the N-terminal and C-terminal fragments after this processing are It is reported to be inactive (see Non-Patent Documents 2 and 3).
これまでに、血清又は血漿中のＦＧＦ－２３の免疫学的測定法が報告されており（特許文献２，３；非特許文献４参照）、免疫学的測定法に基づく測定キットも市販されている［Human Intact FGF-23 ELISA Kit(Immutopics社)、Human FGF-23 ELISA Kit(Merck Millipore社)、FGF-23測定試薬（カイノス社）］。
So far, immunological measurement methods for FGF-23 in serum or plasma have been reported (see Patent Documents 2 and 3; Non-Patent Document 4), and measurement kits based on the immunological measurement methods are also commercially available. [Human Intact FGF-23 ELISA Kit (Immutopics), Human FGF-23 ELISA Kit (Merck Millipore), FGF-23 measuring reagent (Kinos)].
ＦＧＦ－２３は非常に不安定であり、ＦＧＦ－２３を含有する水性媒体を容器内で保存すると、水性媒体中のＦＧＦ－２３濃度が徐々に減少する。そのため、正確な検量線の作成が困難となると共に、検量線に基づく試料中のＦＧＦ－２３の正確な測定が困難になるという問題がある。
FGF-23 is very unstable, and when an aqueous medium containing FGF-23 is stored in a container, the concentration of FGF-23 in the aqueous medium gradually decreases. Therefore, there are problems that it is difficult to prepare an accurate calibration curve and that it is difficult to accurately measure FGF-23 in a sample based on the calibration curve.
本発明の目的は、試料中のＦＧＦ－２３の正確な測定を可能とする、ＦＧＦ－２３の安定化剤、ＦＧＦ－２３の安定化方法、ＦＧＦ－２３の保存方法、及び、ＦＧＦ－２３測定用試薬を提供することにある。
An object of the present invention is to provide an FGF-23 stabilizer, an FGF-23 stabilization method, an FGF-23 storage method, and an FGF-23 measurement, which enable accurate measurement of FGF-23 in a sample. It is to provide a reagent for use.
本発明の発明者は、上記課題を解決すべく鋭意検討し、ポリオキシエチレン系の非イオン性界面活性剤によりＦＧＦ－２３が安定化される、という知見を見出して、本発明を完成させた。すなわち、本発明は、下記［１］～［１７］に関する。
The inventor of the present invention diligently studied to solve the above problems, and found the finding that FGF-23 is stabilized by a polyoxyethylene-based nonionic surfactant, and completed the present invention. . That is, the present invention relates to the following [1] to [17].
［１］ 有効成分としてポリオキシエチレン系の非イオン性界面活性剤を含有することを特徴とする、ＦＧＦ－２３の安定化剤。
［２］ 前記ポリオキシエチレン系の非イオン性界面活性剤が、ポリオキシエチレンソルビタン脂肪酸エステル、ポリオキシエチレンアルキルエーテル、ポリオキシエチレンアルキルフェニルエーテル、及び、ポリオキシエチレンアルキルアミンからなる群より選ばれる少なくとも１種の非イオン性界面活性剤である、前記［１］に記載の安定化剤。
［３］ さらにアルブミンを含有する、前記［１］又は［２］に記載の安定化剤。
［４］ 前記アルブミンがウシ血清アルブミンである、前記［３］に記載の安定化剤。 [1] A stabilizer for FGF-23, which contains a polyoxyethylene-based nonionic surfactant as an active ingredient.
[2] The polyoxyethylene nonionic surfactant is selected from the group consisting of polyoxyethylene sorbitan fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene alkylphenyl ether, and polyoxyethylene alkylamine. The stabilizer according to [1] above, which is at least one nonionic surfactant.
[3] The stabilizer according to [1] or [2], further containing albumin.
[4] The stabilizer according to [3], wherein the albumin is bovine serum albumin.
［２］ 前記ポリオキシエチレン系の非イオン性界面活性剤が、ポリオキシエチレンソルビタン脂肪酸エステル、ポリオキシエチレンアルキルエーテル、ポリオキシエチレンアルキルフェニルエーテル、及び、ポリオキシエチレンアルキルアミンからなる群より選ばれる少なくとも１種の非イオン性界面活性剤である、前記［１］に記載の安定化剤。
［３］ さらにアルブミンを含有する、前記［１］又は［２］に記載の安定化剤。
［４］ 前記アルブミンがウシ血清アルブミンである、前記［３］に記載の安定化剤。 [1] A stabilizer for FGF-23, which contains a polyoxyethylene-based nonionic surfactant as an active ingredient.
[2] The polyoxyethylene nonionic surfactant is selected from the group consisting of polyoxyethylene sorbitan fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene alkylphenyl ether, and polyoxyethylene alkylamine. The stabilizer according to [1] above, which is at least one nonionic surfactant.
[3] The stabilizer according to [1] or [2], further containing albumin.
[4] The stabilizer according to [3], wherein the albumin is bovine serum albumin.
［５］ ＦＧＦ－２３を含有する試料を、容器中で、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体と共存させることを特徴とする、ＦＧＦ－２３の安定化方法。
［６］ 前記ポリオキシエチレン系の非イオン性界面活性剤が、ポリオキシエチレンソルビタン脂肪酸エステル、ポリオキシエチレンアルキルエーテル、ポリオキシエチレンアルキルフェニルエーテル、及び、ポリオキシエチレンアルキルアミンからなる群より選ばれる、少なくとも１種の非イオン性界面活性剤である、前記［５］に記載の安定化方法。
［７］ ＦＧＦ－２３を含有する試料を、さらにアルブミンと共存させる、前記［５］又は［６］に記載の安定化方法。
［８］ 前記アルブミンがウシ血清アルブミンである、前記［７］記載の安定化方法。
［９］ 前記容器がプラスチック製容器である、前記［５］～［８］のいずれか一つに記載の安定化方法。
［１０］ 前記プラスチック製容器が、ポリエチレン製容器、ポリプロピレン製容器、及び、ポリスチレン製容器からなる群より選ばれる容器である、前記［９］に記載の安定化方法。 [5] A method for stabilizing FGF-23, wherein a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
[6] The polyoxyethylene nonionic surfactant is selected from the group consisting of polyoxyethylene sorbitan fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene alkylphenyl ether, and polyoxyethylene alkylamine. The stabilization method according to [5], wherein the stabilization method is at least one nonionic surfactant.
[7] The stabilization method according to [5] or [6] above, wherein a sample containing FGF-23 is further allowed to coexist with albumin.
[8] The stabilization method according to [7], wherein the albumin is bovine serum albumin.
[9] The stabilization method according to any one of [5] to [8], wherein the container is a plastic container.
[10] The stabilization method according to [9], wherein the plastic container is a container selected from the group consisting of a polyethylene container, a polypropylene container, and a polystyrene container.
［６］ 前記ポリオキシエチレン系の非イオン性界面活性剤が、ポリオキシエチレンソルビタン脂肪酸エステル、ポリオキシエチレンアルキルエーテル、ポリオキシエチレンアルキルフェニルエーテル、及び、ポリオキシエチレンアルキルアミンからなる群より選ばれる、少なくとも１種の非イオン性界面活性剤である、前記［５］に記載の安定化方法。
［７］ ＦＧＦ－２３を含有する試料を、さらにアルブミンと共存させる、前記［５］又は［６］に記載の安定化方法。
［８］ 前記アルブミンがウシ血清アルブミンである、前記［７］記載の安定化方法。
［９］ 前記容器がプラスチック製容器である、前記［５］～［８］のいずれか一つに記載の安定化方法。
［１０］ 前記プラスチック製容器が、ポリエチレン製容器、ポリプロピレン製容器、及び、ポリスチレン製容器からなる群より選ばれる容器である、前記［９］に記載の安定化方法。 [5] A method for stabilizing FGF-23, wherein a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
[6] The polyoxyethylene nonionic surfactant is selected from the group consisting of polyoxyethylene sorbitan fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene alkylphenyl ether, and polyoxyethylene alkylamine. The stabilization method according to [5], wherein the stabilization method is at least one nonionic surfactant.
[7] The stabilization method according to [5] or [6] above, wherein a sample containing FGF-23 is further allowed to coexist with albumin.
[8] The stabilization method according to [7], wherein the albumin is bovine serum albumin.
[9] The stabilization method according to any one of [5] to [8], wherein the container is a plastic container.
[10] The stabilization method according to [9], wherein the plastic container is a container selected from the group consisting of a polyethylene container, a polypropylene container, and a polystyrene container.
［１１］ ＦＧＦ－２３を含有する試料を、容器中で、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体と共存させることを特徴とする、ＦＧＦ－２３の保存方法。
［１２］ 前記ポリオキシエチレン系の非イオン性界面活性剤が、ポリオキシエチレンソルビタン脂肪酸エステル、ポリオキシエチレンアルキルエーテル、ポリオキシエチレンアルキルフェニルエーテル、及び、ポリオキシエチレンアルキルアミンからなる群より選ばれる、少なくとも１種の非イオン性界面活性剤である、前記［１１］に記載の保存方法。
［１３］ ＦＧＦ－２３を含有する試料を、さらにアルブミンと共存させる、［１１］又は［１２］に記載の保存方法。
［１４］ 前記アルブミンがウシ血清アルブミンである、前記［１３］に記載の保存方法。
［１５］ 前記容器がプラスチック製容器である、前記［１１］～［１４］のいずれか一つに記載の保存方法。
［１６］ プラスチック製容器が、ポリエチレン製容器、ポリプロピレン製容器、及び、ポリスチレン製容器からなる群より選ばれる何れかの容器である、前記［１５］に記載の保存方法。 [11] A method for storing FGF-23, wherein a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
[12] The polyoxyethylene-based nonionic surfactant is selected from the group consisting of polyoxyethylene sorbitan fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene alkylphenyl ether, and polyoxyethylene alkylamine. The storage method according to [11] above, which is at least one nonionic surfactant.
[13] The storage method according to [11] or [12], wherein a sample containing FGF-23 is further allowed to coexist with albumin.
[14] The storage method according to [13], wherein the albumin is bovine serum albumin.
[15] The storage method according to any one of [11] to [14], wherein the container is a plastic container.
[16] The storage method according to [15], wherein the plastic container is any container selected from the group consisting of a polyethylene container, a polypropylene container, and a polystyrene container.
［１２］ 前記ポリオキシエチレン系の非イオン性界面活性剤が、ポリオキシエチレンソルビタン脂肪酸エステル、ポリオキシエチレンアルキルエーテル、ポリオキシエチレンアルキルフェニルエーテル、及び、ポリオキシエチレンアルキルアミンからなる群より選ばれる、少なくとも１種の非イオン性界面活性剤である、前記［１１］に記載の保存方法。
［１３］ ＦＧＦ－２３を含有する試料を、さらにアルブミンと共存させる、［１１］又は［１２］に記載の保存方法。
［１４］ 前記アルブミンがウシ血清アルブミンである、前記［１３］に記載の保存方法。
［１５］ 前記容器がプラスチック製容器である、前記［１１］～［１４］のいずれか一つに記載の保存方法。
［１６］ プラスチック製容器が、ポリエチレン製容器、ポリプロピレン製容器、及び、ポリスチレン製容器からなる群より選ばれる何れかの容器である、前記［１５］に記載の保存方法。 [11] A method for storing FGF-23, wherein a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
[12] The polyoxyethylene-based nonionic surfactant is selected from the group consisting of polyoxyethylene sorbitan fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene alkylphenyl ether, and polyoxyethylene alkylamine. The storage method according to [11] above, which is at least one nonionic surfactant.
[13] The storage method according to [11] or [12], wherein a sample containing FGF-23 is further allowed to coexist with albumin.
[14] The storage method according to [13], wherein the albumin is bovine serum albumin.
[15] The storage method according to any one of [11] to [14], wherein the container is a plastic container.
[16] The storage method according to [15], wherein the plastic container is any container selected from the group consisting of a polyethylene container, a polypropylene container, and a polystyrene container.
［１７］ 前記［１］～［４］のいずれか一つに記載のＦＧＦ－２３の安定化剤を含有することを特徴とする、ＦＧＦ－２３測定用試薬。
[17] A reagent for measuring FGF-23, comprising the stabilizer for FGF-23 according to any one of [1] to [4].
本発明により、試料中のＦＧＦ－２３の正確な測定を可能とする、ＦＧＦ－２３の安定化剤、ＦＧＦ－２３の安定化方法、ＦＧＦ－２３の保存方法、及び、ＦＧＦ－２３測定用試薬が提供される。
According to the present invention, an FGF-23 stabilizer, an FGF-23 stabilization method, an FGF-23 storage method, and an FGF-23 measurement reagent that enable accurate measurement of FGF-23 in a sample Is provided.
（ＦＧＦ－２３の安定化剤）
本発明のＦＧＦ－２３の安定化剤は、有効成分としてポリオキシエチレン系の非イオン性界面活性剤を含有する。また、本発明のＦＧＦ－２３の安定化剤は、ポリオキシエチレン系の非イオン性界面活性剤に加え、アルブミンを含有してもよい。本発明のＦＧＦ－２３の安定化剤は、容器内に収容されたＦＧＦ－２３水溶液におけるＦＧＦ－２３を安定に保持する効果を有する。ここで、「ＦＧＦ－２３を安定に保持する」とは、ＦＧＦ－２３水溶液を容器内で保存している間、容器内に収容されているＦＧＦ－２３水溶液におけるＦＧＦ－２３濃度がほぼ一定に維持されることを意味する。容器内に収容されているＦＧＦ－２３水溶液におけるＦＧＦ－２３濃度の測定方法は、ＦＧＦ－２３を正確に測定できる方法であれば、如何なる方法でもよく、例えば免疫学的測定法による公知のＦＧＦ－２３の測定方法等を挙げることができる。免疫学的測定法としては、後述の免疫学的測定法等が挙げられる。 (Stabilizer for FGF-23)
The stabilizer for FGF-23 of the present invention contains a polyoxyethylene nonionic surfactant as an active ingredient. Further, the FGF-23 stabilizer of the present invention may contain albumin in addition to the polyoxyethylene nonionic surfactant. The FGF-23 stabilizer of the present invention has an effect of stably holding FGF-23 in an aqueous FGF-23 solution contained in a container. Here, “stable FGF-23” means that the FGF-23 concentration in the FGF-23 aqueous solution contained in the container is substantially constant while the FGF-23 aqueous solution is stored in the container. It means that it is maintained. The method for measuring the FGF-23 concentration in the FGF-23 aqueous solution contained in the container may be any method as long as it can accurately measure FGF-23. For example, a well-known FGF-based immunoassay method may be used. 23 measurement methods and the like. Examples of the immunological measurement method include immunological measurement methods described later.
本発明のＦＧＦ－２３の安定化剤は、有効成分としてポリオキシエチレン系の非イオン性界面活性剤を含有する。また、本発明のＦＧＦ－２３の安定化剤は、ポリオキシエチレン系の非イオン性界面活性剤に加え、アルブミンを含有してもよい。本発明のＦＧＦ－２３の安定化剤は、容器内に収容されたＦＧＦ－２３水溶液におけるＦＧＦ－２３を安定に保持する効果を有する。ここで、「ＦＧＦ－２３を安定に保持する」とは、ＦＧＦ－２３水溶液を容器内で保存している間、容器内に収容されているＦＧＦ－２３水溶液におけるＦＧＦ－２３濃度がほぼ一定に維持されることを意味する。容器内に収容されているＦＧＦ－２３水溶液におけるＦＧＦ－２３濃度の測定方法は、ＦＧＦ－２３を正確に測定できる方法であれば、如何なる方法でもよく、例えば免疫学的測定法による公知のＦＧＦ－２３の測定方法等を挙げることができる。免疫学的測定法としては、後述の免疫学的測定法等が挙げられる。 (Stabilizer for FGF-23)
The stabilizer for FGF-23 of the present invention contains a polyoxyethylene nonionic surfactant as an active ingredient. Further, the FGF-23 stabilizer of the present invention may contain albumin in addition to the polyoxyethylene nonionic surfactant. The FGF-23 stabilizer of the present invention has an effect of stably holding FGF-23 in an aqueous FGF-23 solution contained in a container. Here, “stable FGF-23” means that the FGF-23 concentration in the FGF-23 aqueous solution contained in the container is substantially constant while the FGF-23 aqueous solution is stored in the container. It means that it is maintained. The method for measuring the FGF-23 concentration in the FGF-23 aqueous solution contained in the container may be any method as long as it can accurately measure FGF-23. For example, a well-known FGF-based immunoassay method may be used. 23 measurement methods and the like. Examples of the immunological measurement method include immunological measurement methods described later.
本発明の安定化剤により安定化されるＦＧＦ－２３は、遺伝子組換え方法により製造されたものでも、生体試料から採取されたものであってもよい。遺伝子組換え方法によりＦＧＦ－２３を製造する場合に用いられる遺伝子組換え方法としては、例えば宿主として大腸菌、動物細胞を用いて得られた形質転換体にＦＧＦ－２３を産生させる方法等が挙げられる。生体試料からＦＧＦ－２３を採取する方法としては、例えばＦＧＦ－２３が含まれる可能性がある血液、血漿、血清等の生体試料から、ＦＧＦ－２３を分離、精製する方法等が挙げられる。また、公知の方法（例えば、WO2012/029837参照）を用いて、ＦＧＦ－２３を製造することもできる。
FGF-23 stabilized by the stabilizer of the present invention may be produced by a gene recombination method or collected from a biological sample. Examples of the gene recombination method used for producing FGF-23 by a gene recombination method include a method of producing FGF-23 in a transformant obtained using Escherichia coli as an host and animal cells. . As a method for collecting FGF-23 from a biological sample, for example, a method for separating and purifying FGF-23 from a biological sample such as blood, plasma, or serum that may contain FGF-23. In addition, FGF-23 can also be produced using a known method (for example, see WO2012 / 029837).
ＦＧＦ－２３水溶液とは、ＦＧＦ－２３が水性媒体中に溶解されている溶液である。水性媒体としては、ＦＧＦ－２３を安定に保持し得る水性媒体であれば特に制限はなく、例えば脱イオン水、蒸留水、緩衝液等が挙げられ、これらの中では緩衝液が好ましい。
The FGF-23 aqueous solution is a solution in which FGF-23 is dissolved in an aqueous medium. The aqueous medium is not particularly limited as long as it is capable of stably holding FGF-23, and examples thereof include deionized water, distilled water, and a buffer solution. Among these, a buffer solution is preferable.
緩衝液に用いる緩衝剤としては、例えばトリス（ヒドロキシメチル）アミノメタン緩衝剤、リン酸緩衝剤、ホウ酸緩衝剤、グッドの緩衝剤等が挙げられる。グッドの緩衝剤としては、例えば2-モルホリノエタンスルホン酸（MES）、ビス（2-ヒドロキシエチル）イミノトリス（ヒドロキシメチル）メタン(Bis-Tris)、N-（2-アセトアミド）イミノ二酢酸(ADA)、ピペラジン-N,N’-ビス（2-エタンスルホン酸）(PIPES)、N-（2-アセトアミド）-2-アミノエタンスルホン酸(ACES)、3-モルホリノ-2-ヒドロキシプロパンスルホン酸(MOPSO)、N,N-ビス（2-ヒドロキシエチル）-2-アミノエタンスルホン酸(BES)、3-モルホリノプロパンスルホン酸(MOPS)、N-〔トリス（ヒドロキシメチル）メチル〕-2-アミノエタンスルホン酸(TES)、2-〔4-（2-ヒドロキシエチル）-1-ピペラジニル〕エタンスルホン酸(HEPES)、3-〔N,N-ビス（2-ヒドロキシエチル）アミノ〕-2-ヒドロキシプロパンスルホン酸(DIPSO)、N-〔トリス（ヒドロキシメチル）メチル〕-2-ヒドロキシ-3-アミノプロパンスルホン酸(TAPSO)、ピペラジン-N,N’-ビス（2-ヒドロキシプロパンスルホン酸）(POPSO)、3-〔4-（2-ヒドロキシエチル）-1-ピペラジニル〕-2-ヒドロキシプロパンスルホン酸(HEPPSO)、3-〔4-（2-ヒドロキシエチル）-1-ピペラジニル〕プロパンスルホン酸[(H)EPPS]、N-〔トリス（ヒドロキシメチル）〕グリシン(Tricine)、N,N-ビス（2-ヒドロキシエチル）グリシン(Bicine)、N-トリス（ヒドロキシメチル）メチル-3-アミノプロパンスルホン酸(TAPS)、N-シクロヘキシル-2-アミノエタンスルホン酸(CHES)、N-シクロヘキシル-3-アミノ-2-ヒドロキシプロパンスルホン酸(CAPSO)、N-シクロヘキシル-3-アミノプロパンスルホン酸(CAPS)等が挙げられる。
Examples of the buffer used in the buffer include tris (hydroxymethyl) aminomethane buffer, phosphate buffer, borate buffer, Good's buffer, and the like. Good buffering agents include, for example, 2-morpholinoethanesulfonic acid (MES), bis (2-hydroxyethyl) iminotris (hydroxymethyl) methane (Bis-Tris), N- (2-acetamido) iminodiacetic acid (ADA) Piperazine-N, N'-bis (2-ethanesulfonic acid) (PIPES), N- (2-acetamido) -2-aminoethanesulfonic acid (ACES), 3-morpholino-2-hydroxypropanesulfonic acid (MOPSO) ), N, N-bis (2-hydroxyethyl) -2-aminoethanesulfonic acid (BES), 3-morpholinopropanesulfonic acid (MOPS), N- [tris (hydroxymethyl) methyl] -2-aminoethanesulfone Acid (TES), 2- [4- (2-hydroxyethyl) -1-piperazinyl] ethanesulfonic acid (HEPES), 3- [N, N-bis (2-hydroxyethyl) amino] -2-hydroxypropanesulfone Acid (DIPSO), N- [Tris (hydroxymethyl) methyl] -2-hydroxy -3-aminopropanesulfonic acid (TAPSO), piperazine-N, N'-bis (2-hydroxypropanesulfonic acid) (POPSO), 3- [4- (2-hydroxyethyl) -1-piperazinyl] -2- Hydroxypropanesulfonic acid (HEPPSO), 3- [4- (2-hydroxyethyl) -1-piperazinyl] propanesulfonic acid [(H) EPPS], N- [Tris (hydroxymethyl)] glycine (Tricine), N, N-bis (2-hydroxyethyl) glycine (Bicine), N-tris (hydroxymethyl) methyl-3-aminopropanesulfonic acid (TAPS), N-cyclohexyl-2-aminoethanesulfonic acid (CHES), N-cyclohexyl -3-amino-2-hydroxypropanesulfonic acid (CAPSO), N-cyclohexyl-3-aminopropanesulfonic acid (CAPS), and the like.
本発明における容器とは、ＦＧＦ－２３水溶液を収容する容器であり、ＦＧＦ－２３水溶液を保存するために用いられる容器である。本発明における容器としては、例えばプラスチック製容器等が挙げられる。プラスチック製容器とは、ＦＧＦ－２３含有試料と接触する表面がプラスチックからなる容器を意味し、容器全体がプラスチックからなる容器のみならず、ＦＧＦ－２３含有試料と接触する表面のみがプラスチックからなる容器も包含される。プラスチック製容器としては、例えばポリエチレン製容器、ポリプロピレン製容器、ポリスチレン製容器、ポリ塩化ビニル製容器、ポリウレタン製容器、ポリテトラフルオロエチレン製容器、ポリエチレンテレフタレート製容器、アクリル樹脂製容器、ポリカーボネート製容器等が挙げられ、ポリエチレン製容器、ポリプロピレン製容器、及び、ポリスチレン製容器が好ましい。
The container in the present invention is a container for storing an FGF-23 aqueous solution, and is a container used for storing the FGF-23 aqueous solution. Examples of the container in the present invention include a plastic container. The plastic container means a container whose surface that comes into contact with the FGF-23-containing sample is made of plastic, and is not only a container whose whole container is made of plastic, but is a container whose only surface that comes into contact with the FGF-23-containing sample is made of plastic. Are also included. Examples of the plastic container include a polyethylene container, a polypropylene container, a polystyrene container, a polyvinyl chloride container, a polyurethane container, a polytetrafluoroethylene container, a polyethylene terephthalate container, an acrylic resin container, and a polycarbonate container. A polyethylene container, a polypropylene container, and a polystyrene container are preferable.
本発明におけるポリオキシエチレン系の非イオン性界面活性剤は、ポリオキシエチレン構造を有する非イオン性界面活性剤であれば特に制限はなく、例えばポリオキシエチレンソルビタン脂肪酸エステル（以下、ＰＯＥソルビタン脂肪酸エステルという）、ポリオキシエチレンアルキルエーテル（以下、ＰＯＥアルキルエーテルという）、ポリオキシエチレンアルキルフェニルエーテル（以下、ＰＯＥアルキルフェニルエーテルという）、ポリオキシエチレンアルキルアミン（以下、ＰＯＥアルキルアミンという）、ポリオキシエチレン・ポリオキシプロピレン共重合体（以下、ＰＯＥ・ＰＯＰ共重合体という）、ポリオキシエチレン・ポリオキシプロピレンアルキルエーテル（以下、ＰＯＥ・ＰＯＰアルキルエーテルという）、ポリオキシエチレン・ポリオキシプロピレンアルキルフェニルエーテル（以下、ＰＯＥ・ＰＯＰアルキルフェニルエーテルという）等が挙げられ、これらの中では、ＰＯＥソルビタン脂肪酸エステル、ＰＯＥアルキルエーテル、ＰＯＥアルキルフェニルエーテル、ＰＯＥアルキルアミンが好ましい。本発明において、ポリオキシエチレン系の非イオン性界面活性剤は１種類のみを用いることも、２種類以上を組み合わせて用いることもできる。また、本発明において、ポリオキシエチレン系の非イオン性界面活性剤として、市販品を用いることもできる。
The polyoxyethylene nonionic surfactant in the present invention is not particularly limited as long as it is a nonionic surfactant having a polyoxyethylene structure. For example, polyoxyethylene sorbitan fatty acid ester (hereinafter referred to as POE sorbitan fatty acid ester). Polyoxyethylene alkyl ether (hereinafter referred to as POE alkyl ether), polyoxyethylene alkyl phenyl ether (hereinafter referred to as POE alkyl phenyl ether), polyoxyethylene alkyl amine (hereinafter referred to as POE alkyl amine), polyoxyethylene Polyoxypropylene copolymer (hereinafter referred to as POE / POP copolymer), polyoxyethylene / polyoxypropylene alkyl ether (hereinafter referred to as POE / POP alkyl ether), polyoxyethylene Ethylene-polyoxypropylene alkylphenyl ether (hereinafter, POE-called POP alkyl phenyl ether), and the like. Among these, POE sorbitan fatty acid esters, POE alkyl ethers, POE alkylphenyl ethers, POE alkyl amines are preferred. In the present invention, the polyoxyethylene nonionic surfactant can be used alone or in combination of two or more. In the present invention, commercially available products can also be used as the polyoxyethylene-based nonionic surfactant.
ＰＯＥソルビタン脂肪酸エステルにおける脂肪酸としては、例えば炭素数８～２４の飽和又は不飽和脂肪酸が挙げられ、これらの中では炭素数１２～１８の飽和又は不飽和脂肪酸が好ましい。炭素数８～２４の飽和又は不飽和脂肪酸としては、例えばオクタン酸、デカン酸、ラウリン酸、ミリスチン酸、パルミチン酸、ステアリン酸、オレイン酸、リノール酸、リノレン酸、エイコサトリエン酸、アラキドン酸、イコサン酸、エイコサテトラエン酸、エイコサペンタエン酸、ドコサン酸、ドコサヘキサエン酸、テトラドコサン酸、テトラコサペンタエン酸等が挙げられる。炭素数１２～１８の飽和又は不飽和脂肪酸としては、例えばラウリン酸、ミリスチン酸、パルミチン酸、ステアリン酸、オレイン酸、リノール酸、リノレン酸等が挙げられる。ＰＯＥソルビタン脂肪酸エステルとしては、例えばポリオキシエチレンソルビタンモノラウレート、ポリオキシエチレンソルビタンモノオレエート、ポリオキシエチレンソルビタンモノステアレート、ポリオキシエチレンソルビタンモノパルミテート等が挙げられる。ＰＯＥソルビタン脂肪酸エステルの市販品としては、例えばpolysorbate 80 (Tween 80)（ポリオキシエチレンソルビタンモノオレエート）、polysorbate 40 (Tween 40)（ポリオキシエチレンソルビタンモノパルミテート）、polysorbate 60 (Tween 60)（ポリオキシエチレンソルビタンモノステアレート）、polysorbate 20 (Tween 20)（ポリオキシエチレンソルビタンモノラウレート）（以上、Sigma-Aldrich社製）、BLAUNON OT-21（ポリオキシエチレンソルビタンモノオレエート）（青木油脂工業社製）等が挙げられる。
Examples of the fatty acid in the POE sorbitan fatty acid ester include saturated or unsaturated fatty acids having 8 to 24 carbon atoms, and among these, saturated or unsaturated fatty acids having 12 to 18 carbon atoms are preferable. Examples of saturated or unsaturated fatty acids having 8 to 24 carbon atoms include octanoic acid, decanoic acid, lauric acid, myristic acid, palmitic acid, stearic acid, oleic acid, linoleic acid, linolenic acid, eicosatrienoic acid, arachidonic acid, Examples include icosanoic acid, eicosatetraenoic acid, eicosapentaenoic acid, docosanic acid, docosahexaenoic acid, tetradocosanoic acid, tetracosapentaenoic acid, and the like. Examples of the saturated or unsaturated fatty acid having 12 to 18 carbon atoms include lauric acid, myristic acid, palmitic acid, stearic acid, oleic acid, linoleic acid, and linolenic acid. Examples of the POE sorbitan fatty acid ester include polyoxyethylene sorbitan monolaurate, polyoxyethylene sorbitan monooleate, polyoxyethylene sorbitan monostearate, polyoxyethylene sorbitan monopalmitate and the like. Examples of commercially available POE sorbitan fatty acid esters include polysorbate 80 (Tween 80) (polyoxyethylene sorbitan monooleate), polysorbate 40 (Tween 40) (polyoxyethylene sorbitan monopalmitate), polysorbate 60 (Tween 60) ( Polyoxyethylene sorbitan monostearate), polysorbate 20 (Tween 20) (polyoxyethylene sorbitan monolaurate) (above, Sigma-Aldrich), BLAUNON OT-21 (polyoxyethylene sorbitan monooleate) (Aoki oil) Manufactured by Kogyo Co., Ltd.).
ＰＯＥアルキルエーテルにおけるアルキルとしては、例えば炭素数８～２４のアルキルが挙げられ、これらの中では炭素数１０～２０のアルキルが好ましい。炭素数８～２４のアルキルとしては、例えばオクチル、イソオクチル、ノニル、デシル、イソデシル、ウンデシル、ドデシル（ラウリル）、トリデシル、テトラデシル（ミリスチル）、ペンタデシル、ヘキサデシル（セチル）、ヘプタデシル、オクタデシル（ステアリル）、オレイル、ノナデシル、イコシル、ヘネイコシル、ドコシル（ベヘニル）、トリコシル、テトラコシル等が挙げられる。炭素数１０～２０のアルキルとしては、例えばデシル、イソデシル、ウンデシル、ドデシル（ラウリル）、トリデシル、テトラデシル（ミリスチル）、ペンタデシル、ヘキサデシル（セチル）、ヘプタデシル、オクタデシル（ステアリル）、オレイル、ノナデシル、イコシル等が挙げられる。ＰＯＥアルキルエーテルの市販品としては、例えばBrij35（ポリオキシエチレンラウリルエーテル）、Brij99（ポリオキシエチレンオレイルエーテル）、Brij56（ポリオキシエチレンセチルエーテル）、Brij78（ポリオキシエチレンステアリルエーテル）（以上、Sigma-Aldrich社製）、ニッコールBC-23（ポリオキシエチレンセチルエーテル）（日光ケミカルズ社製）、ノニオンP-210、ノニオンP-213（以上、ポリオキシエチレンセチルエーテル）、ノニオンK-220（ポリオキシエチレンラウリルエーテル）、ノニオンE-205、ノニオンE-215、ノニオンE-230（以上、ポリオキシエチレンオレイルエーテル）、ノニオンS-215、ノニオンS-220（以上、ポリオキシエチレンステアリルエーテル）（以上、日油社製）、エマルゲン120（ポリオキシエチレンラウリルエーテル）、エマルゲン220（ポリオキシエチレンセチルエーテル）、エマルゲン320P（ポリオキシエチレンステアリルエーテル）、エマルゲン420（ポリオキシエチレンオレイルエーテル）（以上、花王社製）、アデカトールLA-875、アデカトールLA-975（以上、ポリオキシエチレンラウリルエーテル）（以上、ＡＤＥＫＡ社製）、エマルミンNL-100、エマルミンNL-110、エマルミンLS-80、エマルミンLS-90、（以上、ポリオキシエチレンラウリルエーテル）（以上、三洋化成工業社製）、BLAUNON EL-1509、BLAUNON EL-1512P、BLAUNON EL-1515、BLAUNON EL-1521（以上、ポリオキシエチレンラウリルエーテル）、BLAUNON CH-310、BLAUNON CH-310L、BLAUNON CH-313（以上、ポリオキシエチレンセチルエーテル）、BLAUNON SR-711（ポリオキシエチレンステアリルエーテル）（以上、青木油脂工業社製）、ノイゲンSD-70、ノイゲンSD-80、ノイゲンSD-110、ノイゲンSD-150（以上、ポリオキシエチレンイソデシルエーテル）、ノイゲンTDS-80、ノイゲンTDS-100、ノイゲンTDS-200D（以上、ポリオキシエチレントリデシルエーテル）（第一工業製薬社製）等が挙げられる。
Examples of the alkyl in the POE alkyl ether include alkyl having 8 to 24 carbon atoms, and among these, alkyl having 10 to 20 carbon atoms is preferable. Examples of the alkyl having 8 to 24 carbon atoms include octyl, isooctyl, nonyl, decyl, isodecyl, undecyl, dodecyl (lauryl), tridecyl, tetradecyl (myristyl), pentadecyl, hexadecyl (cetyl), heptadecyl, octadecyl (stearyl), oleyl , Nonadecyl, icosyl, heneicosyl, docosyl (behenyl), tricosyl, tetracosyl and the like. Examples of the alkyl having 10 to 20 carbon atoms include decyl, isodecyl, undecyl, dodecyl (lauryl), tridecyl, tetradecyl (myristyl), pentadecyl, hexadecyl (cetyl), heptadecyl, octadecyl (stearyl), oleyl, nonadecyl, icosyl and the like. Can be mentioned. Commercially available POE alkyl ethers include, for example, Brij35 (polyoxyethylene lauryl ether), Brij99 (polyoxyethylene oleyl ether), Brij56 (polyoxyethylene cetyl ether), Brij78 (polyoxyethylene stearyl ether) (above, Sigma- Aldrich), Nikkor BC-23 (polyoxyethylene cetyl ether) (Nikko Chemicals), nonion P-210, nonion P-213 (above, polyoxyethylene cetyl ether), nonion K-220 (polyoxyethylene) Lauryl ether), nonion E-205, nonion E-215, nonion E-230 (more, polyoxyethylene oleyl ether), nonion S-215, nonion S-220 (more, polyoxyethylene stearyl ether) (more, day Oil company), Emulgen 120 (polyoxyethylene lauryl ether), Margen 220 (polyoxyethylene cetyl ether), Emulgen 320P (polyoxyethylene stearyl ether), Emargen 420 (polyoxyethylene oleyl ether) (above, manufactured by Kao Corporation), Adekatol LA-875, Adekatol LA-975 (above, Poly Oxyethylene lauryl ether) (above, made by ADEKA), Emalmin NL-100, Emalmine NL-110, Emalmine LS-80, Emalmine LS-90, (above, polyoxyethylene lauryl ether) (above, made by Sanyo Chemical Industries) ), BLAUNON EL-1509, BLAUNON EL-1512P, BLAUNON EL-1515, BLAUNON EL-1521 (above, polyoxyethylene lauryl ether), BLAUNON CH-310, BLAUNON CH-310L, BLAUNON CH-313 (above, polyoxy Ethylene cetyl ether), BLAUNON SR-711 (polyoxyethylene stearyl ether) (above, manufactured by Aoki Yushi Kogyo Co., Ltd.), Gen SD-70, Neugen SD-80, Neugen SD-110, Neugen SD-150 (above, polyoxyethylene isodecyl ether), Neugen TDS-80, Neugen TDS-100, Neugen TDS-200D (above, polyoxyethylene Tridecyl ether) (Daiichi Kogyo Seiyaku Co., Ltd.).
ＰＯＥアルキルフェニルエーテルにおけるアルキルとしては、例えば炭素数８～９のアルキル等が挙げられる。炭素数８～９のアルキルとしては、例えばオクチル、ノニル等が挙げられる。ＰＯＥアルキルフェニルエーテルの市販品としては、例えばトリトンX-100（Sigma-Aldrich社製）（ポリオキシエチレンオクチルフェニルエーテル）、エマルゲン810（ポリオキシエチレンオクチルフェニルエーテル）、エマルゲン910、エマルゲン930（以上、ポリオキシエチレンノニルフェニルエーテル）（以上、花王社製）、ニッコールOP-10（ポリオキシエチレンオクチルフェニルエーテル）、ニッコールNP-10（ポリオキシエチレンノニルフェニルエーテル）（以上、日光ケミカルズ社製）、ノニオンHS-210、ノニオンHS-215、ノニオンHS-220（以上、ポリオキシエチレンオクチルフェニルエーテル）、ノニオンNS-210、ノニオンNS-215、ノニオンNS-220（以上、ポリオキシエチレンノニルフェニルエーテル）（以上、日油社製）、BLAUNON NK-810（青木油脂工業社製）等が挙げられる。
Examples of the alkyl in the POE alkyl phenyl ether include alkyl having 8 to 9 carbon atoms. Examples of the alkyl having 8 to 9 carbon atoms include octyl and nonyl. Examples of commercially available POE alkylphenyl ethers include Triton X-100 (manufactured by Sigma-Aldrich) (polyoxyethylene octylphenyl ether), Emulgen 810 (polyoxyethylene octylphenyl ether), Emulgen 910, Emulgen 930 (above, Polyoxyethylene nonylphenyl ether) (above, manufactured by Kao Corporation), Nikkor OP-10 (polyoxyethylene octylphenyl ether), Nikkor NP-10 (polyoxyethylene nonylphenyl ether) (above, manufactured by Nikko Chemicals), Nonion HS-210, Nonion HS-215, Nonion HS-220 (above, polyoxyethylene octylphenyl ether), Nonion NS-210, Nonion NS-215, Nonion NS-220 (above, polyoxyethylene nonylphenyl ether) (above , Made by NOF Corporation), BLAUNON NK-810 (made by Aoki Oil & Fat Co., Ltd.) ) And the like.
ＰＯＥアルキルアミンにおけるアルキルとしては、例えば炭素数８～２４のアルキルが挙げられ、これらの中では炭素数１０～２０のアルキルが好ましい。炭素数８～２４のアルキルとしては、例えばオクチル、イソオクチル、ノニル、デシル、イソデシル、ウンデシル、ドデシル（ラウリル）、トリデシル、テトラデシル（ミリスチル）、ペンタデシル、ヘキサデシル（セチル）、ヘプタデシル、オクタデシル（ステアリル）、オレイル、ノナデシル、イコシル、ヘネイコシル、ドコシル（ベヘニル）、トリコシル、テトラコシル等が挙げられる。炭素数１０～２０のアルキルとしては、例えばデシル、イソデシル、ウンデシル、ドデシル（ラウリル）、トリデシル、テトラデシル（ミリスチル）、ペンタデシル、ヘキサデシル（セチル）、ヘプタデシル、オクタデシル（ステアリル）、オレイル、ノナデシル、イコシル等が挙げられる。ＰＯＥアルキルアミンの市販品としては、例えばナイミーンL-207（ポリオキシエチレンラウリルアミン）、ナイミーンS-210、ナイミーンS-215、ナイミーンS-220（以上、ポリオキシエチレンステアリルアミン）、ナイミーンT2-210、ナイミーンT2-230［以上、ポリオキシエチレンアルキル（牛脂）アミン］、ナイミーンF-215［ポリオキシエチレンアルキル（ヤシ脂）アミン］（以上、日油社製）、BLAUNON L-210、BLAUNON L-230（以上、ポリオキシエチレンラウリルアミン）、BLAUNON S-210、BLAUNON S-215、BLAUNON S-220、BLAUNON S-230（以上、ポリオキシエチレンステアリルアミン）、BLAUNON S-210T、BLAUNON S-215T、BLAUNON S-220T、BLAUNON S-230T［以上、ポリオキシエチレンアルキル（牛脂）アミン］（以上、青木油脂工業社製）、ニューコールOD420（ポリオキシエチレンステアリルアミン）（日本乳化剤社製）等が挙げられる。
Examples of the alkyl in the POE alkylamine include alkyl having 8 to 24 carbon atoms, and among these, alkyl having 10 to 20 carbon atoms is preferable. Examples of the alkyl having 8 to 24 carbon atoms include octyl, isooctyl, nonyl, decyl, isodecyl, undecyl, dodecyl (lauryl), tridecyl, tetradecyl (myristyl), pentadecyl, hexadecyl (cetyl), heptadecyl, octadecyl (stearyl), oleyl , Nonadecyl, icosyl, heneicosyl, docosyl (behenyl), tricosyl, tetracosyl and the like. Examples of the alkyl having 10 to 20 carbon atoms include decyl, isodecyl, undecyl, dodecyl (lauryl), tridecyl, tetradecyl (myristyl), pentadecyl, hexadecyl (cetyl), heptadecyl, octadecyl (stearyl), oleyl, nonadecyl, icosyl and the like. Can be mentioned. Commercially available POE alkylamines include, for example, Neimine L-207 (polyoxyethylene laurylamine), Neimine S-210, Naimine S-215, Naimine S-220 (above, polyoxyethylene stearylamine), Naimine T2-210 , Naimine T2-230 [above, polyoxyethylene alkyl (beef tallow) amine], Naimine F-215 [polyoxyethylene alkyl (coconut) amine] (above, NOF Corporation), BLAUNON L-210, BLAUNON L- 230 (above, polyoxyethylene laurylamine), BLAUNON S-210, BLAUNON S-215, BLAUNON S-220, BLAUNON S-230 (above, polyoxyethylene stearylamine), BLAUNON S-210T, BLAUNON S-215T, BLAUNON S-220T, BLAUNON S-230T [above, polyoxyethylene alkyl (beef tallow) amine] (above, manufactured by Aoki Oil & Fat Co., Ltd.), New Coal OD420 (polyoxyethylene steer) Triethanolamine) (made in Japan emulsifier Co., Ltd.), and the like.
ＰＯＥ・ＰＯＰ共重合体の市販品としては、例えばプロノン201、プロノン204、プロノン208（以上、日油社製）等が挙げられる。
Examples of commercially available POE / POP copolymers include Pronon 201, Pronon 204, and Pronon 208 (above, NOF Corporation).
ＰＯＥ・ＰＯＰアルキルエーテルにおけるアルキルとしては、例えば炭素数８～２４のアルキルが挙げられ、これらの中では炭素数１０～２０のアルキルが好ましい。炭素数８～２４のアルキルとしては、例えばオクチル、イソオクチル、ノニル、デシル、イソデシル、ウンデシル、ドデシル（ラウリル）、トリデシル、テトラデシル（ミリスチル）、ペンタデシル、ヘキサデシル（セチル）、ヘプタデシル、オクタデシル（ステアリル）、オレイル、ノナデシル、イコシル、ヘネイコシル、ドコシル（ベヘニル）、トリコシル、テトラコシル等が挙げられる。炭素数１０～２０のアルキルとしては、例えばデシル、イソデシル、ウンデシル、ドデシル（ラウリル）、トリデシル、テトラデシル（ミリスチル）、ペンタデシル、ヘキサデシル（セチル）、ヘプタデシル、オクタデシル（ステアリル）、オレイル、ノナデシル、イコシル等が挙げられる。ＰＯＥ・ＰＯＰアルキルエーテルの市販品としては、例えばファインサーフIDEP-802、ワンダサーフID-90（以上、ポリオキシエチレンポリオキシプロピレンイソデシルエーテル）、ワンダサーフRL-80、ワンダサーフRL-100、ワンダサーフRL-140（以上、ポリオキシエチレンポリオキシプロピレンラウリルエーテル）（以上、青木油脂工業社製）等が挙げられる。
Examples of the alkyl in the POE / POP alkyl ether include alkyl having 8 to 24 carbon atoms, and among these, alkyl having 10 to 20 carbon atoms is preferable. Examples of the alkyl having 8 to 24 carbon atoms include octyl, isooctyl, nonyl, decyl, isodecyl, undecyl, dodecyl (lauryl), tridecyl, tetradecyl (myristyl), pentadecyl, hexadecyl (cetyl), heptadecyl, octadecyl (stearyl), oleyl , Nonadecyl, icosyl, heneicosyl, docosyl (behenyl), tricosyl, tetracosyl and the like. Examples of the alkyl having 10 to 20 carbon atoms include decyl, isodecyl, undecyl, dodecyl (lauryl), tridecyl, tetradecyl (myristyl), pentadecyl, hexadecyl (cetyl), heptadecyl, octadecyl (stearyl), oleyl, nonadecyl, icosyl and the like. Can be mentioned. Examples of commercially available POE / POP alkyl ethers include Fine Surf IDEP-802, Wanda Surf ID-90 (above, polyoxyethylene polyoxypropylene isodecyl ether), Wanda Surf RL-80, Wanda Surf RL-100, Wanda Surf RL-140 (above, polyoxyethylene polyoxypropylene lauryl ether) (above, manufactured by Aoki Yushi Kogyo Co., Ltd.) and the like.
ＰＯＥ・ＰＯＰアルキルフェニルエーテルの市販品としては、例えばエマルゲンL40（ポリオキシエチレンポリオキシプロピレンオクチルフェニルエーテル）（花王社製）等が挙げられる。
Examples of commercially available POE / POP alkylphenyl ethers include Emulgen L40 (polyoxyethylene polyoxypropylene octylphenyl ether) (manufactured by Kao Corporation).
本発明におけるポリオキシエチレン系の非イオン性界面活性のHLB価（Hydrophile-Lipophile Balance価）は、ＦＧＦ－２３を安定化し得るHLB価であれば特に制限はなく、通常12～20であり、12.5～19が好ましく、13～18がより好ましい。使用されるポリオキシエチレン系の非イオン性界面活性剤の濃度は、ＦＧＦ－２３を安定化し得る濃度であれば特に制限はなく、通常0.00001～10w/v%であり、0.00005～1w/v%が好ましい。
The HLB value (Hydrophile-Lipophile Balance value) of the polyoxyethylene-based nonionic surfactant in the present invention is not particularly limited as long as it is an HLB value capable of stabilizing FGF-23, and is usually 12 to 20, 12.5 ~ 19 are preferred, and 13-18 are more preferred. The concentration of the polyoxyethylene-based nonionic surfactant to be used is not particularly limited as long as it is a concentration capable of stabilizing FGF-23, and is usually 0.00001 to 10 w / v%, Is preferred.
本発明におけるアルブミンとしては、例えばウシ、ウマ、ヒツジ、ヒト、マウス、ウサギ由来のアルブミン等が挙げられ、これらの中ではウシ血清アルブミン（ＢＳＡ）が好ましい。また、遺伝子工学的な手法により製造されたアルブミンも用いることができる。本発明においては、２種類以上のアルブミンを組み合わせて使用することもできる。使用されるアルブミンの濃度は、ＦＧＦ－２３を安定化し得る濃度であれば特に制限はなく、通常0.01～5w/v%であり、0.05～1w/v%が好ましい。
Examples of albumin in the present invention include bovine, horse, sheep, human, mouse, and rabbit-derived albumin, and among these, bovine serum albumin (BSA) is preferable. Also, albumin produced by genetic engineering techniques can be used. In the present invention, two or more types of albumin can be used in combination. The concentration of albumin used is not particularly limited as long as it can stabilize FGF-23, and is usually 0.01 to 5 w / v%, preferably 0.05 to 1 w / v%.
本発明の安定化剤には、キレート剤、塩類、防腐剤等を含有してもよい。キレート剤としては、例えばエチレンジアミン四酢酸(EDTA)等が挙げられる。塩類としては、例えば塩化ナトリウム、塩化カリウム、硫酸ナトリウム、硫酸カリウム、硝酸ナトリウム、硝酸カリウム、塩化マグネシウム、塩化カルシウム、硫酸マグネシウム、硫酸カルシウム、硝酸マグネシウム、硝酸カルシウム等が挙げられる。防腐剤としては、例えばアジ化ナトリウム、バイオエース等が挙げられる。
The stabilizer of the present invention may contain a chelating agent, salts, preservatives and the like. Examples of the chelating agent include ethylenediaminetetraacetic acid (EDTA). Examples of the salts include sodium chloride, potassium chloride, sodium sulfate, potassium sulfate, sodium nitrate, potassium nitrate, magnesium chloride, calcium chloride, magnesium sulfate, calcium sulfate, magnesium nitrate, and calcium nitrate. Examples of the preservative include sodium azide and bioacet.
本発明のＦＧＦ－２３安定化剤によるＦＧＦ－２３の安定性は以下の方法により評価することができる。
The stability of FGF-23 by the FGF-23 stabilizer of the present invention can be evaluated by the following method.
先ず、容器中に、ＦＧＦ－２３を含有する溶液Ｘを調製する。調製直後の溶液Ｘ中のＦＧＦ－２３濃度［ＣＸ（調製直後）］を決定する。次に、この溶液Ｘを4℃で7日間保存し、この保存後の溶液Ｘ中のＦＧＦ－２３濃度［ＣＸ（保存後）］を決定する。決定したＣＸ（調製直後）及びＣＸ（保存後）を用いて、以下の式（Ｉ）より、溶液Ｘに対するＦＧＦ－２３の残存率（％）を算出する。
First, a solution X containing FGF-23 is prepared in a container. FGF-23 concentration in the solution X immediately after preparation [CX (immediately after preparation) is determined. Next, the solution X is stored at 4 ° C. for 7 days, and the FGF-23 concentration [CX (after storage) ] in the solution X after the storage is determined. Using the determined CX (immediately after preparation) and CX (after storage) , the residual ratio (%) of FGF-23 to the solution X is calculated from the following formula (I).
溶液Ｘに対するＦＧＦ－２３の残存率（％）は、１００（％）に近い程、ＦＧＦ－２３が安定に保持されていることを意味する。すなわち、ＦＧＦ－２３が安定化されることを意味する。
The closer the percentage of FGF-23 remaining to solution X (%) is to 100 (%), the more stable FGF-23 is retained. That is, it means that FGF-23 is stabilized.
上記のＦＧＦ－２３濃度は、ＦＧＦ－２３を正確に測定できる方法であれば、如何なる方法によっても決定することができ、例えば免疫学的測定法による公知のＦＧＦ－２３測定方法を用いて決定することができる。免疫学的測定法としては、測定原理に基づいて、サンドイッチ法、競合法等が挙げられ、また、検出法に基づいて、吸光度法、化学発光法、蛍光法、放射免疫測定法等が挙げられる。従って、ＦＧＦ－２３濃度は、ＦＧＦ－２３濃度に対応する吸光度、発光強度、蛍光強度、放射線強度等を用いて決定することができる。免疫学的測定法による公知のＦＧＦ－２３測定方法には、市販のＦＧＦ－２３測定キットを用いる方法も含まれる。市販のＦＧＦ－２３測定キットとしては、例えば、Human Intact FGF-23 ELISA Kit(Immutopics社)、Human FGF-23 ELISA Kit(Merck Millipore社)、FGF-23測定試薬（カイノス社）等が挙げられる。
The concentration of FGF-23 can be determined by any method as long as it can accurately measure FGF-23. For example, the FGF-23 concentration is determined by using a known FGF-23 measurement method by immunoassay. be able to. Examples of the immunological measurement method include a sandwich method and a competitive method based on the measurement principle, and examples include an absorbance method, a chemiluminescence method, a fluorescence method, and a radioimmunoassay method based on the detection method. . Therefore, the FGF-23 concentration can be determined using the absorbance, emission intensity, fluorescence intensity, radiation intensity, etc. corresponding to the FGF-23 concentration. Known FGF-23 measurement methods by immunoassay include a method using a commercially available FGF-23 measurement kit. Examples of commercially available FGF-23 measurement kits include Human Intact FGF-23 ELISA KIT (Immutopics), Human FGF-23 ELISA KIT (Merck Millipore), and FGF-23 measurement reagent (Kinos).
（ＦＧＦ－２３の安定化方法）
本発明のＦＧＦ－２３の安定化方法は、ＦＧＦ－２３を含有する試料を、容器中で、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体と共存させることを特徴とする方法である。当該安定化方法の具体的態様を以下に記す。
・安定化方法１
ＦＧＦ－２３を含有する試料を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法２
ポリオキシエチレン系の非イオン性界面活性剤を含む容器へ、ＦＧＦ－２３を含有する試料を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法３
ＦＧＦ－２３を含有する試料を含有する水性媒体を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法４
ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料を添加する工程を有する、ＦＧＦ－２３の安定化方法。
また、ＦＧＦ－２３の安定化方法は、ＦＧＦ－２３を含有する試料を、容器中で、ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含有する水性媒体と共存させることを特徴とする方法である。当該安定化方法の具体的態様を以下に記す。
・安定化方法５
ＦＧＦ－２３を含有する試料を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法６
ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を含む容器へ、アルブミンを含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法７
ＦＧＦ－２３を含有する試料及びアルブミンを含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法８
ポリオキシエチレン系の非イオン性界面活性剤を含む容器へ、ＦＧＦ－２３を含有する試料及びアルブミンを含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法９
ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含む容器へ、ＦＧＦ－２３を含有する試料を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１０
アルブミンを含む容器へ、ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１１
ＦＧＦ－２３を含有する試料を含有する水性媒体を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１２
ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を含む容器へ、アルブミンを添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１３
ＦＧＦ－２３を含有する試料及びアルブミンを含有する水性媒体を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１４
ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料及びアルブミンを添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１５
ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１６
アルブミンを含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を添加する工程を有する、ＦＧＦ－２３の安定化方法。
ＦＧＦ－２３を含有する試料としては、ＦＧＦ－２３を含有する試料であれば特に制限はなく、例えばＦＧＦ－２３を含有する生体試料や、ＦＧＦ－２３標準液等が挙げられる。生体試料としては、例えば血液、血清、血漿、尿、唾液、髄液、糞便等が挙げられる。ＦＧＦ－２３標準液は、既知濃度のＦＧＦ－２３を含有する水溶液であって、生体から取得された試料中のＦＧＦ－２３の定量に必要な検量線の作成等に用いられる。ＦＧＦ－２３標準液に使用されるＦＧＦ－２３としては、例えば前述のＦＧＦ－２３が挙げられる。 (Method for stabilizing FGF-23)
The method for stabilizing FGF-23 of the present invention is characterized in that a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant. It is. Specific embodiments of the stabilization method will be described below.
・ Stabilization method 1
A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a container containing a sample containing FGF-23.
・ Stabilization method 2
A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a sample containing FGF-23 to a container containing a polyoxyethylene-based nonionic surfactant.
・ Stabilization method 3
A method for stabilizing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant to a container containing an aqueous medium containing a sample containing FGF-23.
・ Stabilization method 4
A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
The method for stabilizing FGF-23 is characterized in that a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene nonionic surfactant and albumin. Is the method. Specific embodiments of the stabilization method will be described below.
・ Stabilization method 5
A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene nonionic surfactant and albumin to a container containing a sample containing FGF-23.
・ Stabilization method 6
A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing albumin to a container containing FGF-23 and a container containing a polyoxyethylene-based nonionic surfactant.
・ Stabilization method 7
A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a sample containing FGF-23 and a container containing albumin.
・ Stabilization method 8
A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 and an aqueous medium containing albumin to a container containing a polyoxyethylene-based nonionic surfactant.
・ Stabilization method 9
A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a sample containing FGF-23 to a container containing a polyoxyethylene-based nonionic surfactant and albumin.
・ Stabilization method 10
A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 and an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a container containing albumin.
・ Stabilization method 11
A method for stabilizing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant and albumin to a container containing an aqueous medium containing a sample containing FGF-23.
・ Stabilization method 12
A method for stabilizing FGF-23, comprising a step of adding albumin to a container containing a sample containing FGF-23 and an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
・ Stabilization method 13
A method for stabilizing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant to a container containing a sample containing FGF-23 and an aqueous medium containing albumin.
・ Stabilization method 14
A method for stabilizing FGF-23, comprising the step of adding a sample containing FGF-23 and albumin to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
・ Stabilization method 15
A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 to a container containing an aqueous medium containing polyoxyethylene-based nonionic surfactant and albumin.
・ Stabilization method 16
A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 and a polyoxyethylene-based nonionic surfactant to a container containing an aqueous medium containing albumin.
The sample containing FGF-23 is not particularly limited as long as it is a sample containing FGF-23, and examples thereof include biological samples containing FGF-23 and FGF-23 standard solutions. Examples of the biological sample include blood, serum, plasma, urine, saliva, spinal fluid, stool, and the like. The FGF-23 standard solution is an aqueous solution containing a known concentration of FGF-23, and is used for preparing a calibration curve necessary for quantifying FGF-23 in a sample obtained from a living body. Examples of FGF-23 used in the FGF-23 standard solution include the aforementioned FGF-23.
本発明のＦＧＦ－２３の安定化方法は、ＦＧＦ－２３を含有する試料を、容器中で、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体と共存させることを特徴とする方法である。当該安定化方法の具体的態様を以下に記す。
・安定化方法１
ＦＧＦ－２３を含有する試料を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法２
ポリオキシエチレン系の非イオン性界面活性剤を含む容器へ、ＦＧＦ－２３を含有する試料を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法３
ＦＧＦ－２３を含有する試料を含有する水性媒体を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法４
ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料を添加する工程を有する、ＦＧＦ－２３の安定化方法。
また、ＦＧＦ－２３の安定化方法は、ＦＧＦ－２３を含有する試料を、容器中で、ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含有する水性媒体と共存させることを特徴とする方法である。当該安定化方法の具体的態様を以下に記す。
・安定化方法５
ＦＧＦ－２３を含有する試料を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法６
ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を含む容器へ、アルブミンを含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法７
ＦＧＦ－２３を含有する試料及びアルブミンを含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法８
ポリオキシエチレン系の非イオン性界面活性剤を含む容器へ、ＦＧＦ－２３を含有する試料及びアルブミンを含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法９
ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含む容器へ、ＦＧＦ－２３を含有する試料を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１０
アルブミンを含む容器へ、ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１１
ＦＧＦ－２３を含有する試料を含有する水性媒体を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１２
ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を含む容器へ、アルブミンを添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１３
ＦＧＦ－２３を含有する試料及びアルブミンを含有する水性媒体を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１４
ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料及びアルブミンを添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１５
ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料を添加する工程を有する、ＦＧＦ－２３の安定化方法。
・安定化方法１６
アルブミンを含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を添加する工程を有する、ＦＧＦ－２３の安定化方法。
ＦＧＦ－２３を含有する試料としては、ＦＧＦ－２３を含有する試料であれば特に制限はなく、例えばＦＧＦ－２３を含有する生体試料や、ＦＧＦ－２３標準液等が挙げられる。生体試料としては、例えば血液、血清、血漿、尿、唾液、髄液、糞便等が挙げられる。ＦＧＦ－２３標準液は、既知濃度のＦＧＦ－２３を含有する水溶液であって、生体から取得された試料中のＦＧＦ－２３の定量に必要な検量線の作成等に用いられる。ＦＧＦ－２３標準液に使用されるＦＧＦ－２３としては、例えば前述のＦＧＦ－２３が挙げられる。 (Method for stabilizing FGF-23)
The method for stabilizing FGF-23 of the present invention is characterized in that a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant. It is. Specific embodiments of the stabilization method will be described below.
・ Stabilization method 1
A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a container containing a sample containing FGF-23.
・ Stabilization method 2
A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a sample containing FGF-23 to a container containing a polyoxyethylene-based nonionic surfactant.
・ Stabilization method 3
A method for stabilizing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant to a container containing an aqueous medium containing a sample containing FGF-23.
・ Stabilization method 4
A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
The method for stabilizing FGF-23 is characterized in that a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene nonionic surfactant and albumin. Is the method. Specific embodiments of the stabilization method will be described below.
・ Stabilization method 5
A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene nonionic surfactant and albumin to a container containing a sample containing FGF-23.
・ Stabilization method 6
A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing albumin to a container containing FGF-23 and a container containing a polyoxyethylene-based nonionic surfactant.
・ Stabilization method 7
A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a sample containing FGF-23 and a container containing albumin.
・ Stabilization method 8
A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 and an aqueous medium containing albumin to a container containing a polyoxyethylene-based nonionic surfactant.
・ Stabilization method 9
A method for stabilizing FGF-23, comprising a step of adding an aqueous medium containing a sample containing FGF-23 to a container containing a polyoxyethylene-based nonionic surfactant and albumin.
・ Stabilization method 10
A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 and an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a container containing albumin.
・ Stabilization method 11
A method for stabilizing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant and albumin to a container containing an aqueous medium containing a sample containing FGF-23.
・ Stabilization method 12
A method for stabilizing FGF-23, comprising a step of adding albumin to a container containing a sample containing FGF-23 and an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
・ Stabilization method 13
A method for stabilizing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant to a container containing a sample containing FGF-23 and an aqueous medium containing albumin.
・ Stabilization method 14
A method for stabilizing FGF-23, comprising the step of adding a sample containing FGF-23 and albumin to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
・ Stabilization method 15
A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 to a container containing an aqueous medium containing polyoxyethylene-based nonionic surfactant and albumin.
・ Stabilization method 16
A method for stabilizing FGF-23, comprising a step of adding a sample containing FGF-23 and a polyoxyethylene-based nonionic surfactant to a container containing an aqueous medium containing albumin.
The sample containing FGF-23 is not particularly limited as long as it is a sample containing FGF-23, and examples thereof include biological samples containing FGF-23 and FGF-23 standard solutions. Examples of the biological sample include blood, serum, plasma, urine, saliva, spinal fluid, stool, and the like. The FGF-23 standard solution is an aqueous solution containing a known concentration of FGF-23, and is used for preparing a calibration curve necessary for quantifying FGF-23 in a sample obtained from a living body. Examples of FGF-23 used in the FGF-23 standard solution include the aforementioned FGF-23.
本発明のＦＧＦ－２３の安定化方法において用いられる容器、ポリオキシエチレン系の非イオン性界面活性剤、及び、アルブミンとしては、例えば、前述の容器、ポリオキシエチレン系の非イオン性界面活性剤、及び、アルブミンがそれぞれ挙げられる。
Containers used in the method for stabilizing FGF-23 of the present invention, polyoxyethylene-based nonionic surfactants, and albumin include, for example, the aforementioned containers, polyoxyethylene-based nonionic surfactants And albumin.
本発明のＦＧＦ－２３の安定化方法において、生体試料中のＦＧＦ－２３の含有量は、通常5～10,000 pg/mLであり、検量線の作成も通常、この濃度範囲のＦＧＦ－２３標準液を用いて行われる。本発明のＦＧＦ－２３の安定化方法において使用されるポリオキシエチレン系の非イオン性界面活性剤とその濃度、及び、アルブミンとその濃度としては、例えば、前述のポリオキシエチレン系の非イオン性界面活性剤とその濃度、及び、アルブミンとその濃度がそれぞれ挙げられる。
In the method for stabilizing FGF-23 of the present invention, the content of FGF-23 in the biological sample is usually 5 to 10,000 μpg / mL, and the calibration curve is usually prepared in the FGF-23 standard solution in this concentration range. It is done using. Examples of the polyoxyethylene-based nonionic surfactant and its concentration, and albumin and its concentration used in the method for stabilizing FGF-23 of the present invention include the aforementioned polyoxyethylene-based nonionic surfactants. Surfactant and its concentration, and albumin and its concentration are mentioned, respectively.
本発明のＦＧＦ－２３の安定化方法における安定化期間は、ＦＧＦ－２３が安定に保持される期間であれば特に制限はなく、通常、7日～1年間である。また、本発明のＦＧＦ－２３の安定化方法において、ＦＧＦ－２３を含有する試料を、容器中で、ポリオキシエチレン系の非イオン性界面活性剤、又は、ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンと共存させる温度は、ＦＧＦ－２３が安定に保持される温度であれば特に制限はなく、通常、-5～45℃であり、0～30℃が好ましく、2～10℃が特に好ましい。
The stabilization period in the FGF-23 stabilization method of the present invention is not particularly limited as long as FGF-23 is stably held, and is usually 7 days to 1 year. Further, in the method for stabilizing FGF-23 of the present invention, a sample containing FGF-23 is placed in a container in a polyoxyethylene-based nonionic surfactant or a polyoxyethylene-based nonionic interface. The temperature for coexisting with the active agent and albumin is not particularly limited as long as FGF-23 is stably maintained, and is usually -5 to 45 ° C, preferably 0 to 30 ° C, and preferably 2 to 10 ° C. Particularly preferred.
本発明のＦＧＦ－２３の安定化方法においては、ポリオキシエチレン系の非イオン性界面活性剤およびアルブミンの他に、キレート剤、塩類、防腐剤等を、ＦＧＦ－２３水溶液中に添加してもよい。キレート剤、塩類、防腐剤としては、例えば、前述のキレート剤、塩類、防腐剤がそれぞれ挙げられる。
In the method for stabilizing FGF-23 of the present invention, in addition to the polyoxyethylene nonionic surfactant and albumin, chelating agents, salts, preservatives and the like may be added to the FGF-23 aqueous solution. Good. Examples of chelating agents, salts, and preservatives include the aforementioned chelating agents, salts, and preservatives.
（ＦＧＦ－２３の保存方法）
本発明のＦＧＦ－２３の保存方法は、ＦＧＦ－２３を含有する試料を、容器中で、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体と共存させることを特徴とする方法である。当該保存方法の具体的態様を以下に記す。
・保存方法１
ＦＧＦ－２３を含有する試料を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法２
ポリオキシエチレン系の非イオン性界面活性剤を含む容器へ、ＦＧＦ－２３を含有する試料を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法３
ＦＧＦ－２３を含有する試料を含有する水性媒体を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法４
ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料を添加する工程を有する、ＦＧＦ－２３の保存方法。
また、ＦＧＦ－２３の保存方法は、ＦＧＦ－２３を含有する試料を、容器中で、ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含有する水性媒体と共存させることを特徴とする方法である。当該保存方法の具体的態様を以下に記す。
・保存方法５
ＦＧＦ－２３を含有する試料を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法６
ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を含む容器へ、アルブミンを含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法７
ＦＧＦ－２３を含有する試料及びアルブミンを含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法８
ポリオキシエチレン系の非イオン性界面活性剤を含む容器へ、ＦＧＦ－２３を含有する試料及びアルブミンを含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法９
ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含む容器へ、ＦＧＦ－２３を含有する試料を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１０
アルブミンを含む容器へ、ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１１
ＦＧＦ－２３を含有する試料を含有する水性媒体を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１２
ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を含む容器へ、アルブミンを添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１３
ＦＧＦ－２３を含有する試料及びアルブミンを含有する水性媒体を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１４
ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料及びアルブミンを添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１５
ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１６
アルブミンを含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を添加する工程を有する、ＦＧＦ－２３の保存方法。
本発明のＦＧＦ－２３の保存方法に使用されるおけるＦＧＦ－２３とその濃度、ポリオキシエチレン系の非イオン性界面活性剤とその濃度、及び、アルブミンとその濃度としては、例えば、上記のＦＧＦ－２３の安定化剤におけるＦＧＦ－２３とその濃度、ポリオキシエチレン系の非イオン性界面活性剤とその濃度、及び、アルブミンとその濃度がそれぞれ挙げられる。本発明のＦＧＦ－２３の保存方法における保存期間は、ＦＧＦ－２３が安定に保存される期間であれば特に制限はなく、通常、7日～1年間である。また、本発明のＦＧＦ－２３の保存方法における保存温度は、ＦＧＦ－２３が安定に保存される温度であれば特に制限はなく、通常、-5～45℃であり、0～30℃が好ましく、2～10℃が特に好ましい。 (Preservation method of FGF-23)
The method for preserving FGF-23 of the present invention is a method characterized in that a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant. is there. Specific embodiments of the storage method will be described below.
・ Storage method 1
A method for storing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a container containing a sample containing FGF-23.
・ Storage method 2
A method for storing FGF-23, comprising a step of adding an aqueous medium containing a sample containing FGF-23 to a container containing a polyoxyethylene-based nonionic surfactant.
・ Storage method 3
A method for storing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant to a container containing an aqueous medium containing a sample containing FGF-23.
・ Save method 4
A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
In addition, a method for storing FGF-23 is characterized in that a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant and albumin. It is. Specific embodiments of the storage method will be described below.
・ Save method 5
A method for storing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant and an aqueous medium containing albumin to a container containing a sample containing FGF-23.
・ Storage method 6
A method for storing FGF-23, comprising a step of adding an aqueous medium containing albumin to a container containing FGF-23 and a container containing a polyoxyethylene-based nonionic surfactant.
・ Storage method 7
A method for storing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a sample containing FGF-23 and a container containing albumin.
・ Storage method 8
A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 and an aqueous medium containing albumin to a container containing a polyoxyethylene-based nonionic surfactant.
・ Storage method 9
A method for storing FGF-23, comprising a step of adding an aqueous medium containing a sample containing FGF-23 to a container containing a polyoxyethylene-based nonionic surfactant and albumin.
・ Storage method 10
A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 and an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a container containing albumin.
・ Storage method 11
A method for storing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant and albumin to a container containing an aqueous medium containing a sample containing FGF-23.
・ Storage method 12
A method for storing FGF-23, comprising a step of adding albumin to a container containing a sample containing FGF-23 and an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
・ Storage method 13
A method for storing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant to a container containing a sample containing FGF-23 and an aqueous medium containing albumin.
・ Storage method 14
A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 and albumin to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
・ Storage method 15
A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant and albumin.
・ Storage method 16
A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 and a polyoxyethylene-based nonionic surfactant to a container containing an aqueous medium containing albumin.
Examples of FGF-23 and its concentration, polyoxyethylene nonionic surfactant and its concentration, and albumin and its concentration used in the method for storing FGF-23 of the present invention include the above-mentioned FGF. Examples thereof include FGF-23 and its concentration in a stabilizer of −23, polyoxyethylene nonionic surfactant and its concentration, and albumin and its concentration. The storage period in the FGF-23 storage method of the present invention is not particularly limited as long as FGF-23 is stably stored, and is usually 7 days to 1 year. The storage temperature in the method for storing FGF-23 of the present invention is not particularly limited as long as FGF-23 is stably stored, and is usually -5 to 45 ° C, preferably 0 to 30 ° C. 2 to 10 ° C. is particularly preferable.
本発明のＦＧＦ－２３の保存方法は、ＦＧＦ－２３を含有する試料を、容器中で、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体と共存させることを特徴とする方法である。当該保存方法の具体的態様を以下に記す。
・保存方法１
ＦＧＦ－２３を含有する試料を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法２
ポリオキシエチレン系の非イオン性界面活性剤を含む容器へ、ＦＧＦ－２３を含有する試料を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法３
ＦＧＦ－２３を含有する試料を含有する水性媒体を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法４
ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料を添加する工程を有する、ＦＧＦ－２３の保存方法。
また、ＦＧＦ－２３の保存方法は、ＦＧＦ－２３を含有する試料を、容器中で、ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含有する水性媒体と共存させることを特徴とする方法である。当該保存方法の具体的態様を以下に記す。
・保存方法５
ＦＧＦ－２３を含有する試料を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法６
ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を含む容器へ、アルブミンを含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法７
ＦＧＦ－２３を含有する試料及びアルブミンを含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法８
ポリオキシエチレン系の非イオン性界面活性剤を含む容器へ、ＦＧＦ－２３を含有する試料及びアルブミンを含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法９
ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含む容器へ、ＦＧＦ－２３を含有する試料を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１０
アルブミンを含む容器へ、ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１１
ＦＧＦ－２３を含有する試料を含有する水性媒体を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１２
ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を含む容器へ、アルブミンを添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１３
ＦＧＦ－２３を含有する試料及びアルブミンを含有する水性媒体を含む容器へ、ポリオキシエチレン系の非イオン性界面活性剤を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１４
ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料及びアルブミンを添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１５
ポリオキシエチレン系の非イオン性界面活性剤及びアルブミンを含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料を添加する工程を有する、ＦＧＦ－２３の保存方法。
・保存方法１６
アルブミンを含有する水性媒体を含む容器へ、ＦＧＦ－２３を含有する試料及びポリオキシエチレン系の非イオン性界面活性剤を添加する工程を有する、ＦＧＦ－２３の保存方法。
本発明のＦＧＦ－２３の保存方法に使用されるおけるＦＧＦ－２３とその濃度、ポリオキシエチレン系の非イオン性界面活性剤とその濃度、及び、アルブミンとその濃度としては、例えば、上記のＦＧＦ－２３の安定化剤におけるＦＧＦ－２３とその濃度、ポリオキシエチレン系の非イオン性界面活性剤とその濃度、及び、アルブミンとその濃度がそれぞれ挙げられる。本発明のＦＧＦ－２３の保存方法における保存期間は、ＦＧＦ－２３が安定に保存される期間であれば特に制限はなく、通常、7日～1年間である。また、本発明のＦＧＦ－２３の保存方法における保存温度は、ＦＧＦ－２３が安定に保存される温度であれば特に制限はなく、通常、-5～45℃であり、0～30℃が好ましく、2～10℃が特に好ましい。 (Preservation method of FGF-23)
The method for preserving FGF-23 of the present invention is a method characterized in that a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant. is there. Specific embodiments of the storage method will be described below.
・ Storage method 1
A method for storing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a container containing a sample containing FGF-23.
・ Storage method 2
A method for storing FGF-23, comprising a step of adding an aqueous medium containing a sample containing FGF-23 to a container containing a polyoxyethylene-based nonionic surfactant.
・ Storage method 3
A method for storing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant to a container containing an aqueous medium containing a sample containing FGF-23.
・ Save method 4
A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
In addition, a method for storing FGF-23 is characterized in that a sample containing FGF-23 is allowed to coexist in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant and albumin. It is. Specific embodiments of the storage method will be described below.
・ Save method 5
A method for storing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant and an aqueous medium containing albumin to a container containing a sample containing FGF-23.
・ Storage method 6
A method for storing FGF-23, comprising a step of adding an aqueous medium containing albumin to a container containing FGF-23 and a container containing a polyoxyethylene-based nonionic surfactant.
・ Storage method 7
A method for storing FGF-23, comprising a step of adding an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a sample containing FGF-23 and a container containing albumin.
・ Storage method 8
A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 and an aqueous medium containing albumin to a container containing a polyoxyethylene-based nonionic surfactant.
・ Storage method 9
A method for storing FGF-23, comprising a step of adding an aqueous medium containing a sample containing FGF-23 to a container containing a polyoxyethylene-based nonionic surfactant and albumin.
・ Storage method 10
A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 and an aqueous medium containing a polyoxyethylene-based nonionic surfactant to a container containing albumin.
・ Storage method 11
A method for storing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant and albumin to a container containing an aqueous medium containing a sample containing FGF-23.
・ Storage method 12
A method for storing FGF-23, comprising a step of adding albumin to a container containing a sample containing FGF-23 and an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
・ Storage method 13
A method for storing FGF-23, comprising a step of adding a polyoxyethylene-based nonionic surfactant to a container containing a sample containing FGF-23 and an aqueous medium containing albumin.
・ Storage method 14
A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 and albumin to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant.
・ Storage method 15
A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 to a container containing an aqueous medium containing a polyoxyethylene-based nonionic surfactant and albumin.
・ Storage method 16
A method for storing FGF-23, comprising a step of adding a sample containing FGF-23 and a polyoxyethylene-based nonionic surfactant to a container containing an aqueous medium containing albumin.
Examples of FGF-23 and its concentration, polyoxyethylene nonionic surfactant and its concentration, and albumin and its concentration used in the method for storing FGF-23 of the present invention include the above-mentioned FGF. Examples thereof include FGF-23 and its concentration in a stabilizer of −23, polyoxyethylene nonionic surfactant and its concentration, and albumin and its concentration. The storage period in the FGF-23 storage method of the present invention is not particularly limited as long as FGF-23 is stably stored, and is usually 7 days to 1 year. The storage temperature in the method for storing FGF-23 of the present invention is not particularly limited as long as FGF-23 is stably stored, and is usually -5 to 45 ° C, preferably 0 to 30 ° C. 2 to 10 ° C. is particularly preferable.
本発明のＦＧＦ－２３の保存方法においては、ポリオキシエチレン系の非イオン性界面活性剤およびアルブミンの他に、キレート剤、塩類、防腐剤等を、ＦＧＦ－２３水溶液中に添加してもよい。キレート剤、塩類、防腐剤としては、例えば、前述のキレート剤、塩類、防腐剤がそれぞれ挙げられる。
In the FGF-23 storage method of the present invention, in addition to the polyoxyethylene nonionic surfactant and albumin, chelating agents, salts, preservatives, and the like may be added to the FGF-23 aqueous solution. . Examples of chelating agents, salts, and preservatives include the aforementioned chelating agents, salts, and preservatives.
（ＦＧＦ－２３測定用試薬）
本発明のＦＧＦ－２３測定用試薬は、本発明のＦＧＦ－２３の安定化剤を含む。本発明のＦＧＦ－２３測定用試薬は、本発明のＦＧＦ－２３の安定化剤以外の、その他の成分を含むものとすることができる。この場合、本発明のＦＧＦ－２３測定用試薬においては、本発明のＦＧＦ－２３の安定化剤を、その他の成分とは独立した容器に入れて、その他の成分が入れられた容器とは独立した構成要素としてもよいが、他のＦＧＦ－２３測定用試薬に含有させて、同一の容器に入れることもできる。これらの形態の中では、本発明のＦＧＦ－２３の安定化剤は、その他の成分と同一の容器に入れることが好ましい。「その他の成分」としては、例えば試料希釈液、磁性粒子懸濁液、ビオチン結合抗ＦＧＦ－２３抗体含有試薬、アルカリホスファターゼ標識抗ＦＧＦ－２３抗体フラグメント含有試薬、ＦＧＦ－２３測定用標準液等が挙げられるが、本発明のＦＧＦ－２３の安定化剤を試料希釈液及び／又はＦＧＦ－２３測定用標準液に含有させる形態が好ましい。既知濃度のＦＧＦ－２３を含有するＦＧＦ－２３測定用標準液に本発明のＦＧＦ－２３の安定化剤を含有させることにより、標準品中のＦＧＦ－２３が安定化されるため、正確な検量線を作成することができ、正確な検量線に基づく、正確なＦＧＦ－２３の測定が可能となる。 (Reagent for FGF-23 measurement)
The reagent for measuring FGF-23 of the present invention contains the stabilizer for FGF-23 of the present invention. The reagent for measuring FGF-23 of the present invention may contain other components other than the stabilizer for FGF-23 of the present invention. In this case, in the reagent for measuring FGF-23 of the present invention, the stabilizer for FGF-23 of the present invention is placed in a container independent of other components and independent of the container in which other components are placed. However, it may be contained in another FGF-23 measurement reagent and placed in the same container. Among these forms, the FGF-23 stabilizer of the present invention is preferably placed in the same container as the other components. Examples of the “other components” include a sample diluent, a magnetic particle suspension, a biotin-conjugated anti-FGF-23 antibody-containing reagent, an alkaline phosphatase-labeled anti-FGF-23 antibody fragment-containing reagent, and a standard solution for FGF-23 measurement. As an example, a form in which the FGF-23 stabilizer of the present invention is contained in a sample diluent and / or a standard solution for FGF-23 measurement is preferable. By containing the stabilizer for FGF-23 of the present invention in a standard solution for FGF-23 measurement containing a known concentration of FGF-23, FGF-23 in the standard product is stabilized, so that accurate calibration is possible. A line can be created, and an accurate measurement of FGF-23 based on an accurate calibration curve is possible.
本発明のＦＧＦ－２３測定用試薬は、本発明のＦＧＦ－２３の安定化剤を含む。本発明のＦＧＦ－２３測定用試薬は、本発明のＦＧＦ－２３の安定化剤以外の、その他の成分を含むものとすることができる。この場合、本発明のＦＧＦ－２３測定用試薬においては、本発明のＦＧＦ－２３の安定化剤を、その他の成分とは独立した容器に入れて、その他の成分が入れられた容器とは独立した構成要素としてもよいが、他のＦＧＦ－２３測定用試薬に含有させて、同一の容器に入れることもできる。これらの形態の中では、本発明のＦＧＦ－２３の安定化剤は、その他の成分と同一の容器に入れることが好ましい。「その他の成分」としては、例えば試料希釈液、磁性粒子懸濁液、ビオチン結合抗ＦＧＦ－２３抗体含有試薬、アルカリホスファターゼ標識抗ＦＧＦ－２３抗体フラグメント含有試薬、ＦＧＦ－２３測定用標準液等が挙げられるが、本発明のＦＧＦ－２３の安定化剤を試料希釈液及び／又はＦＧＦ－２３測定用標準液に含有させる形態が好ましい。既知濃度のＦＧＦ－２３を含有するＦＧＦ－２３測定用標準液に本発明のＦＧＦ－２３の安定化剤を含有させることにより、標準品中のＦＧＦ－２３が安定化されるため、正確な検量線を作成することができ、正確な検量線に基づく、正確なＦＧＦ－２３の測定が可能となる。 (Reagent for FGF-23 measurement)
The reagent for measuring FGF-23 of the present invention contains the stabilizer for FGF-23 of the present invention. The reagent for measuring FGF-23 of the present invention may contain other components other than the stabilizer for FGF-23 of the present invention. In this case, in the reagent for measuring FGF-23 of the present invention, the stabilizer for FGF-23 of the present invention is placed in a container independent of other components and independent of the container in which other components are placed. However, it may be contained in another FGF-23 measurement reagent and placed in the same container. Among these forms, the FGF-23 stabilizer of the present invention is preferably placed in the same container as the other components. Examples of the “other components” include a sample diluent, a magnetic particle suspension, a biotin-conjugated anti-FGF-23 antibody-containing reagent, an alkaline phosphatase-labeled anti-FGF-23 antibody fragment-containing reagent, and a standard solution for FGF-23 measurement. As an example, a form in which the FGF-23 stabilizer of the present invention is contained in a sample diluent and / or a standard solution for FGF-23 measurement is preferable. By containing the stabilizer for FGF-23 of the present invention in a standard solution for FGF-23 measurement containing a known concentration of FGF-23, FGF-23 in the standard product is stabilized, so that accurate calibration is possible. A line can be created, and an accurate measurement of FGF-23 based on an accurate calibration curve is possible.
以下、実施例により本発明をより詳細に説明するが、これらは本発明の範囲を何ら限定するものではない。尚、本実施例においては、下記メーカーの試薬及び容器を使用した。尚、ＦＧＦ－２３は、特許文献２に記載された方法によって製造したものを使用した。
EXAMPLES Hereinafter, although an Example demonstrates this invention in detail, these do not limit the scope of the present invention at all. In this example, reagents and containers from the following manufacturers were used. Note that FGF-23 manufactured by the method described in Patent Document 2 was used.
Polysorbate 80（ポリオキシエチレンソルビタンモノオレエート；HLB 15.0）（Sigma-Aldrich社製）、Polysorbate 20（ポリオキシエチレンソルビタンモノラウレート；HLB 16.7）（Sigma-Aldrich社製）、Brij 35（ポリオキシエチレンラウリルエーテル；HLB 16.9）（Sigma-Aldrich社製）、Brij 99（ポリオキシエチレンオレイルエーテル；HLB 15.3）（Sigma-Aldrich社製）、Triton X-100（ポリオキシエチレンオクチルフェニルエーテル；HLB 16.9）（Sigma-Aldrich社製）、ナイミーンS-220（ポリオキシエチレンオクタデシルアミン；HLB 15.4）（日油社製）、ニッサンカチオンPB-300（ヘキサデシルトリメチルアンモニウム クロライド）（日油社製）、ニッサンカチオンF2-35R［アルキル（ヤシ）ジメチルベンジルアンモニウム クロライド］（日油社製）、デオキシコール酸ナトリウム（関東化学社製）、パーソフトEP（アルキルエーテル硫酸ナトリウム塩）（日油社製）、CHAPS｛２－［（３－コールアミドプロピル）ジメチルアンモニオ］プロパンスルホネート｝（同仁化学研究所社製）、ニッサンアノンBL（ラウリルジメチルアミノ酢酸ベタイン）（日油社製）、ＢＳＡ（Bovogen Biologicals社製)、塩化ナトリウム（関東化学社製）。
Polysorbate 80 (polyoxyethylene sorbitan monooleate; HLB 15.0) (Sigma-Aldrich), Polysorbate 20 (polyoxyethylene sorbitan monolaurate; HLB 16.7) (Sigma-Aldrich), Brij 35 (polyoxyethylene) Lauryl ether; HLB 16.9) (manufactured by Sigma-Aldrich), Brij 99 (polyoxyethylene oleyl ether; HLB 15.3) (manufactured by Sigma-Aldrich), Triton X-100 (polyoxyethylene octylphenyl ether; HLB 16.9) ( Sigma-Aldrich), Naimine S-220 (polyoxyethylene octadecylamine; HLB 15.4) (manufactured by NOF Corporation), Nissan Cation PB-300 (hexadecyltrimethylammonium chloride) (manufactured by NOF Corporation), Nissan Cation F2 -35R [alkyl (coconut) dimethylbenzylammonium chloride] (manufactured by NOF Corporation), sodium deoxycholate (Manufactured by Kanto Chemical Co., Inc.), persoft EP (alkyl ether sulfate sodium salt) (manufactured by NOF Corporation), CHAPS {2-[(3-cholamidopropyl) dimethylammonio] propanesulfonate} (manufactured by Dojindo Laboratories) ), Nissan Anon BL (Betarine Lauryldimethylaminoacetate) (manufactured by NOF Corporation), BSA (manufactured by Bovogen Biologics), sodium chloride (manufactured by Kanto Chemical Co., Inc.).
容器［ポリプロピレン製(1.5 mL)マイクロテストチューブ］（トーホー社製）。
Container [Polypropylene (1.5 mL) micro test tube] (Toho).
ポリオキシエチレン系非イオン性界面活性剤によるＦＧＦ－２３の安定化
（１）ＦＧＦ－２３溶液（溶液Ａ１～Ａ６；溶液ａ１～ａ６）の調製
以下の組成からなるＦＧＦ－２３溶液（溶液Ａ１～Ａ６；溶液ａ１～ａ６）を調製した。
リン酸緩衝液 10 mmol/L (pH7.4)
ＦＧＦ－２３ 1μg/mL
塩化ナトリウム 140 mmol/L
界面活性剤（第１表参照） 0.0005% Stabilization of FGF-23 by polyoxyethylene nonionic surfactant (1) Preparation of FGF-23 solution (solutions A1 to A6; solutions a1 to a6) FGF-23 solution (solutions A1 to A6) having the following composition A6; Solutions a1 to a6) were prepared.
Phosphate buffer 10 mmol / L (pH7.4)
FGF-23 1μg / mL
Sodium chloride 140 mmol / L
Surfactant (See Table 1) 0.0005%
（１）ＦＧＦ－２３溶液（溶液Ａ１～Ａ６；溶液ａ１～ａ６）の調製
以下の組成からなるＦＧＦ－２３溶液（溶液Ａ１～Ａ６；溶液ａ１～ａ６）を調製した。
リン酸緩衝液 10 mmol/L (pH7.4)
ＦＧＦ－２３ 1μg/mL
塩化ナトリウム 140 mmol/L
界面活性剤（第１表参照） 0.0005% Stabilization of FGF-23 by polyoxyethylene nonionic surfactant (1) Preparation of FGF-23 solution (solutions A1 to A6; solutions a1 to a6) FGF-23 solution (solutions A1 to A6) having the following composition A6; Solutions a1 to a6) were prepared.
Phosphate buffer 10 mmol / L (pH7.4)
FGF-23 1μg / mL
Sodium chloride 140 mmol / L
Surfactant (See Table 1) 0.0005%
（２）ＦＧＦ－２３溶液（溶液ａ０）の調製
以下の組成からなるＦＧＦ－２３溶液（溶液ａ０）を調製した。
リン酸緩衝液 10 mmol/L (pH7.4)
ＦＧＦ－２３ 1μg/mL
塩化ナトリウム 140 mmol/L
溶液ａ０は、界面活性剤が含まれていない以外は、溶液Ａ１～Ａ６、及び、溶液ａ１～ａ６と同じ組成である。 (2) Preparation of FGF-23 solution (solution a0) An FGF-23 solution (solution a0) having the following composition was prepared.
Phosphate buffer 10 mmol / L (pH7.4)
FGF-23 1μg / mL
Sodium chloride 140 mmol / L
The solution a0 has the same composition as the solutions A1 to A6 and the solutions a1 to a6 except that the surfactant is not included.
以下の組成からなるＦＧＦ－２３溶液（溶液ａ０）を調製した。
リン酸緩衝液 10 mmol/L (pH7.4)
ＦＧＦ－２３ 1μg/mL
塩化ナトリウム 140 mmol/L
溶液ａ０は、界面活性剤が含まれていない以外は、溶液Ａ１～Ａ６、及び、溶液ａ１～ａ６と同じ組成である。 (2) Preparation of FGF-23 solution (solution a0) An FGF-23 solution (solution a0) having the following composition was prepared.
Phosphate buffer 10 mmol / L (pH7.4)
FGF-23 1μg / mL
Sodium chloride 140 mmol / L
The solution a0 has the same composition as the solutions A1 to A6 and the solutions a1 to a6 except that the surfactant is not included.
（３）抗原希釈液の調製
以下の組成からなる抗原希釈液を調製した。
リン酸緩衝液 20 mmol/L (pH6.5)
塩化ナトリウム 300 mmol/L
ＢＳＡ 0.2(w/v)% (3) Preparation of antigen dilution liquid An antigen dilution liquid having the following composition was prepared.
Phosphate buffer solution 20 mmol / L (pH6.5)
Sodium chloride 300 mmol / L
BSA 0.2 (w / v)%
以下の組成からなる抗原希釈液を調製した。
リン酸緩衝液 20 mmol/L (pH6.5)
塩化ナトリウム 300 mmol/L
ＢＳＡ 0.2(w/v)% (3) Preparation of antigen dilution liquid An antigen dilution liquid having the following composition was prepared.
Phosphate buffer solution 20 mmol / L (pH6.5)
Sodium chloride 300 mmol / L
BSA 0.2 (w / v)%
（４）ＦＧＦ－２３定量用キット
以下の磁性粒子懸濁液、ビオチン結合抗ＦＧＦ－２３抗体含有試薬、及び、アルカリホスファターゼ標識抗ＦＧＦ－２３抗体フラグメント含有試薬の各構成試薬からなるＦＧＦ－２３定量用キットを調製した。 (4) FGF-23 quantification kit FGF-23 quantification comprising the following magnetic particle suspension, biotin-conjugated anti-FGF-23 antibody-containing reagent, and alkaline phosphatase-labeled anti-FGF-23 antibody fragment-containing reagent. A kit was prepared.
以下の磁性粒子懸濁液、ビオチン結合抗ＦＧＦ－２３抗体含有試薬、及び、アルカリホスファターゼ標識抗ＦＧＦ－２３抗体フラグメント含有試薬の各構成試薬からなるＦＧＦ－２３定量用キットを調製した。 (4) FGF-23 quantification kit FGF-23 quantification comprising the following magnetic particle suspension, biotin-conjugated anti-FGF-23 antibody-containing reagent, and alkaline phosphatase-labeled anti-FGF-23 antibody fragment-containing reagent. A kit was prepared.
＜磁性粒子懸濁液＞
磁性粒子として、ストレプトアビジンが結合した市販の磁性粒子（Dynabeads MyOne Streptavidin；ダイナル社製）を用いて、以下の組成からなる磁性粒子懸濁液を調製した。
ＭＥＳ（ｐＨ６．５） 50 mmol/L
ストレプトアビジン結合磁性粒子 0.75 g/L
ＢＳＡ 0.1%
塩化ナトリウム 0.1 mol/L <Suspension of magnetic particles>
As magnetic particles, commercially available magnetic particles (Dynabeads MyOne Streptavidin; manufactured by Dynal) bound with streptavidin were used to prepare a magnetic particle suspension having the following composition.
MES (pH 6.5) 50 mmol / L
Streptavidin-coupled magnetic particles 0.75 g / L
BSA 0.1%
Sodium chloride 0.1 mol / L
磁性粒子として、ストレプトアビジンが結合した市販の磁性粒子（Dynabeads MyOne Streptavidin；ダイナル社製）を用いて、以下の組成からなる磁性粒子懸濁液を調製した。
ＭＥＳ（ｐＨ６．５） 50 mmol/L
ストレプトアビジン結合磁性粒子 0.75 g/L
ＢＳＡ 0.1%
塩化ナトリウム 0.1 mol/L <Suspension of magnetic particles>
As magnetic particles, commercially available magnetic particles (Dynabeads MyOne Streptavidin; manufactured by Dynal) bound with streptavidin were used to prepare a magnetic particle suspension having the following composition.
MES (pH 6.5) 50 mmol / L
Streptavidin-coupled magnetic particles 0.75 g / L
BSA 0.1%
Sodium chloride 0.1 mol / L
＜ビオチン結合抗ＦＧＦ－２３抗体含有試薬＞
第１抗体として、FERM BP-7838として寄託されたハイブリドーマが生産する抗ＦＧＦ－２３モノクローナル抗体を用いて、当該抗体とNHS－ビオチンとを混合し、３７℃で１時間反応させ、反応後の混合物をセファデックスG-25カラム（ＧＥヘルスサイエンス・ジャパン社製）に供して未反応のNHS－ビオチンを除去し、ビオチン結合抗ＦＧＦ－２３モノクローナル抗体を調製した。得られたビオチン結合抗ＦＧＦ－２３モノクローナル抗体を用いて、以下の組成からなるビオチン結合抗ＦＧＦ－２３抗体含有試薬を調製した。
ＭＥＳ（ｐＨ６．５） 50 mmol/L
抗ＦＧＦ－２３モノクローナル抗体 5μg/mL
ＢＳＡ 0.1%
塩化ナトリウム 0.1 mol/L <Biotin-conjugated anti-FGF-23 antibody-containing reagent>
Using the anti-FGF-23 monoclonal antibody produced by the hybridoma deposited as FERM BP-7838 as the first antibody, the antibody and NHS-biotin are mixed, reacted at 37 ° C. for 1 hour, and the mixture after the reaction Was applied to a Sephadex G-25 column (manufactured by GE Health Science Japan) to remove unreacted NHS-biotin, and a biotin-conjugated anti-FGF-23 monoclonal antibody was prepared. Using the obtained biotin-conjugated anti-FGF-23 monoclonal antibody, a biotin-conjugated anti-FGF-23 antibody-containing reagent having the following composition was prepared.
MES (pH 6.5) 50 mmol / L
Anti-FGF-23 monoclonal antibody 5μg / mL
BSA 0.1%
Sodium chloride 0.1 mol / L
第１抗体として、FERM BP-7838として寄託されたハイブリドーマが生産する抗ＦＧＦ－２３モノクローナル抗体を用いて、当該抗体とNHS－ビオチンとを混合し、３７℃で１時間反応させ、反応後の混合物をセファデックスG-25カラム（ＧＥヘルスサイエンス・ジャパン社製）に供して未反応のNHS－ビオチンを除去し、ビオチン結合抗ＦＧＦ－２３モノクローナル抗体を調製した。得られたビオチン結合抗ＦＧＦ－２３モノクローナル抗体を用いて、以下の組成からなるビオチン結合抗ＦＧＦ－２３抗体含有試薬を調製した。
ＭＥＳ（ｐＨ６．５） 50 mmol/L
抗ＦＧＦ－２３モノクローナル抗体 5μg/mL
ＢＳＡ 0.1%
塩化ナトリウム 0.1 mol/L <Biotin-conjugated anti-FGF-23 antibody-containing reagent>
Using the anti-FGF-23 monoclonal antibody produced by the hybridoma deposited as FERM BP-7838 as the first antibody, the antibody and NHS-biotin are mixed, reacted at 37 ° C. for 1 hour, and the mixture after the reaction Was applied to a Sephadex G-25 column (manufactured by GE Health Science Japan) to remove unreacted NHS-biotin, and a biotin-conjugated anti-FGF-23 monoclonal antibody was prepared. Using the obtained biotin-conjugated anti-FGF-23 monoclonal antibody, a biotin-conjugated anti-FGF-23 antibody-containing reagent having the following composition was prepared.
MES (pH 6.5) 50 mmol / L
Anti-FGF-23 monoclonal antibody 5μg / mL
BSA 0.1%
Sodium chloride 0.1 mol / L
＜アルカリホスファターゼ標識抗ＦＧＦ－２３抗体フラグメント含有試薬＞
第２抗体フラグメントとして、FERM BP-7839として寄託されたハイブリドーマが生産する抗ＦＧＦ－２３モノクローナル抗体をペプシンで消化した後、G3000SWカラム（東ソー社製；口径：21.5 mm；長さ：60 cm）を用いたHPLCシステム（日立製作所社製）でF(ab’)2を分離した。得られたF(ab’)2を２－メルカプトエチルアミン塩酸塩（ナカライテスク社製）で還元した後、G3000SWカラム（東ソー社製；口径：21.5 mm；長さ：60 cm）を用いたHPLCシステム（日立製作所社製）でFab’を分離した。得られたFab’とアルカリホスファターゼとを以下の手順により、マレイミド法によって結合させた。 <Alkaline phosphatase labeled anti-FGF-23 antibody fragment-containing reagent>
As a second antibody fragment, an anti-FGF-23 monoclonal antibody produced by the hybridoma deposited as FERM BP-7839 was digested with pepsin, and then a G3000SW column (manufactured by Tosoh Corporation; aperture: 21.5 mm; length: 60 cm) was used. F (ab ′) 2 was separated using the HPLC system (manufactured by Hitachi, Ltd.). The obtained F (ab ′) 2 was reduced with 2-mercaptoethylamine hydrochloride (manufactured by Nacalai Tesque), and then a HPLC system using a G3000SW column (manufactured by Tosoh Corporation; aperture: 21.5 mm; length: 60 cm) Fab 'was separated by (manufactured by Hitachi, Ltd.). The obtained Fab ′ and alkaline phosphatase were bound by the maleimide method according to the following procedure.
第２抗体フラグメントとして、FERM BP-7839として寄託されたハイブリドーマが生産する抗ＦＧＦ－２３モノクローナル抗体をペプシンで消化した後、G3000SWカラム（東ソー社製；口径：21.5 mm；長さ：60 cm）を用いたHPLCシステム（日立製作所社製）でF(ab’)2を分離した。得られたF(ab’)2を２－メルカプトエチルアミン塩酸塩（ナカライテスク社製）で還元した後、G3000SWカラム（東ソー社製；口径：21.5 mm；長さ：60 cm）を用いたHPLCシステム（日立製作所社製）でFab’を分離した。得られたFab’とアルカリホスファターゼとを以下の手順により、マレイミド法によって結合させた。 <Alkaline phosphatase labeled anti-FGF-23 antibody fragment-containing reagent>
As a second antibody fragment, an anti-FGF-23 monoclonal antibody produced by the hybridoma deposited as FERM BP-7839 was digested with pepsin, and then a G3000SW column (manufactured by Tosoh Corporation; aperture: 21.5 mm; length: 60 cm) was used. F (ab ′) 2 was separated using the HPLC system (manufactured by Hitachi, Ltd.). The obtained F (ab ′) 2 was reduced with 2-mercaptoethylamine hydrochloride (manufactured by Nacalai Tesque), and then a HPLC system using a G3000SW column (manufactured by Tosoh Corporation; aperture: 21.5 mm; length: 60 cm) Fab 'was separated by (manufactured by Hitachi, Ltd.). The obtained Fab ′ and alkaline phosphatase were bound by the maleimide method according to the following procedure.
マレイミド化試薬Sulfo-HMCS（同仁化学研究所社製）を用いて、アルカリホスファターゼをマレイミド化し、反応混合物をセファデックスG-25カラム（ＧＥヘルスサイエンス・ジャパン社製）に供して未反応のSulfo-HMCSを除去し、マレイミド化アルカリホスファターゼを得た。
Using maleimide-forming reagent Sulfo-HMCS (manufactured by Dojindo Laboratories), alkaline phosphatase was maleimidated, and the reaction mixture was subjected to Sephadex G-25 column (GE Health Science Japan) to make unreacted Sulfo- HMCS was removed to obtain maleimidated alkaline phosphatase.
調製したマレイミド化アルカリホスファターゼとFab’とを混合し、アルカリホスファターゼ標識Fab’抗体を作製した。得られたアルカリホスファターゼ標識Fab’抗体を用いて、以下の組成からなるアルカリホスファターゼ標識抗ＦＧＦ－２３抗体フラグメント溶液を調製した。
ＭＥＳ（ｐＨ６．５） 50 mmol/L
アルカリホスファターゼ標識抗ＦＧＦ－２３抗体フラグメント 7μg/mL
ＢＳＡ 0.1%
塩化ナトリウム 0.1 mol/L The prepared maleimidated alkaline phosphatase and Fab ′ were mixed to prepare an alkaline phosphatase labeled Fab ′ antibody. Using the obtained alkaline phosphatase labeled Fab ′ antibody, an alkaline phosphatase labeled anti-FGF-23 antibody fragment solution having the following composition was prepared.
MES (pH 6.5) 50 mmol / L
Alkaline phosphatase-labeled anti-FGF-23 antibody fragment 7 μg / mL
BSA 0.1%
Sodium chloride 0.1 mol / L
ＭＥＳ（ｐＨ６．５） 50 mmol/L
アルカリホスファターゼ標識抗ＦＧＦ－２３抗体フラグメント 7μg/mL
ＢＳＡ 0.1%
塩化ナトリウム 0.1 mol/L The prepared maleimidated alkaline phosphatase and Fab ′ were mixed to prepare an alkaline phosphatase labeled Fab ′ antibody. Using the obtained alkaline phosphatase labeled Fab ′ antibody, an alkaline phosphatase labeled anti-FGF-23 antibody fragment solution having the following composition was prepared.
MES (pH 6.5) 50 mmol / L
Alkaline phosphatase-labeled anti-FGF-23 antibody fragment 7 μg / mL
BSA 0.1%
Sodium chloride 0.1 mol / L
（５）検量線の作成
次の各濃度のＦＧＦ－２３の抗原希釈液溶液を測定用試料として調製した：10,000 pg/mL；3,000 pg/mL；1,000 pg/mL；300 pg/mL；100 pg/mL；50 pg/mL；5 pg/mL。なお、抗原希釈液として、0.2(w/v)％BSA及び300 mmol/L塩化ナトリウムを含む、20 mmol/L リン酸緩衝液(pH 6.5)を使用した。 (5) Preparation of calibration curve Antigen dilution solution of each of the following concentrations of FGF-23 was prepared as a measurement sample: 10,000 pg / mL; 3,000 pg / mL; 1,000 pg / mL; 300 pg / mL; 100 pg / mL; 50 pg / mL; 5 pg / mL. A 20 mmol / L phosphate buffer (pH 6.5) containing 0.2 (w / v)% BSA and 300 mmol / L sodium chloride was used as the antigen dilution.
次の各濃度のＦＧＦ－２３の抗原希釈液溶液を測定用試料として調製した：10,000 pg/mL；3,000 pg/mL；1,000 pg/mL；300 pg/mL；100 pg/mL；50 pg/mL；5 pg/mL。なお、抗原希釈液として、0.2(w/v)％BSA及び300 mmol/L塩化ナトリウムを含む、20 mmol/L リン酸緩衝液(pH 6.5)を使用した。 (5) Preparation of calibration curve Antigen dilution solution of each of the following concentrations of FGF-23 was prepared as a measurement sample: 10,000 pg / mL; 3,000 pg / mL; 1,000 pg / mL; 300 pg / mL; 100 pg / mL; 50 pg / mL; 5 pg / mL. A 20 mmol / L phosphate buffer (pH 6.5) containing 0.2 (w / v)% BSA and 300 mmol / L sodium chloride was used as the antigen dilution.
調製した測定用試料(10μL)を反応キュベットに添加し、次いで、上記（４）で調製した磁性粒子懸濁液、ビオチン結合抗ＦＧＦ－２３抗体溶液、及び、アルカリホスファターゼ標識抗ＦＧＦ－２３抗体フラグメント溶液の各構成試薬30μLずつ加えて撹拌し、37℃で10分間反応させた。反応混合液中の磁性粒子を磁力で集めて、磁性粒子以外の反応液を除去すると共に、洗浄液［0.05%Tween20及び140 mmol/L塩化ナトリウムを含有する10 mmol/L リン酸緩衝液(pH7.4)］で磁性粒子を5回洗浄した。その後、９－（４－クロロフェニルチオホスホリルオキシメチリデン）－１０－メチルアクリダン・二ナトリウム塩を主成分とする発光基質液(100μL)を加えて撹拌し、生成した光の発光強度を測定した。10,000 pg/mLのＦＧＦ－２３の抗原希釈液溶液の代わりに、3,000 pg/mLのＦＧＦ－２３の抗原希釈液溶液、1,000 pg/mLのＦＧＦ－２３の抗原希釈液溶液、300 pg/mLのＦＧＦ－２３の抗原希釈液溶液、100 pg/mLのＦＧＦ－２３の抗原希釈液溶液、50 pg/mLのＦＧＦ－２３の抗原希釈液溶液、5 pg/mLのＦＧＦ－２３の抗原希釈液溶液、及び、上記（３）で調製した抗原希釈液の各溶液を用いて、同様の測定を行い、各溶液に対する発光強度を測定した。測定した発光強度を基に、ＦＧＦ－２３濃度と、発光強度との間の関係を示す検量線を作成した。
The prepared measurement sample (10 μL) was added to the reaction cuvette, and then the magnetic particle suspension prepared in (4) above, the biotin-conjugated anti-FGF-23 antibody solution, and the alkaline phosphatase-labeled anti-FGF-23 antibody fragment 30 μL of each constituent reagent of the solution was added and stirred, and reacted at 37 ° C. for 10 minutes. The magnetic particles in the reaction mixture are collected by magnetic force to remove the reaction liquid other than the magnetic particles, and at the same time, the washing liquid [10% mmol / L phosphate buffer (pH 7.7 containing 0.05% Tween20 and 140% mmol / L sodium chloride). 4)], the magnetic particles were washed five times. Thereafter, a luminescent substrate solution (100 μL) mainly composed of 9- (4-chlorophenylthiophosphoryloxymethylidene) -10-methylacridan disodium salt was added and stirred, and the luminescence intensity of the generated light was measured. . Instead of 10,000 pg / mL FGF-23 antigen dilution solution, 3,000 pg / mL FGF-23 antigen dilution solution, 1,000 pg / mL FGF-23 antigen dilution solution, 300 pg / mL FGF-23 antigen dilution solution, 100 pg / mL FGF-23 antigen dilution solution, 50 pg / mL FGF-23 antigen dilution solution, 5 pg / mL FGF-23 antigen dilution solution And the same measurement was performed using each solution of the antigen dilution liquid prepared in said (3), and the emitted light intensity with respect to each solution was measured. A calibration curve showing the relationship between the FGF-23 concentration and the luminescence intensity was prepared based on the measured luminescence intensity.
（６）調製直後の各ＦＧＦ－２３溶液（溶液Ａ１～Ａ６；溶液ａ０～ａ６）中のＦＧＦ－２３の測定
上記（１）で調製した直後の溶液Ａ１(0.5 mL)を、上記（２）で調製した抗原希釈液で500倍希釈し、測定用試料を調製した。調製した測定用試料を用いて、上記（５）と同様の方法により測定を行い、溶液Ａ１に対する発光強度を測定した。測定した発光強度を上記（５）で作成した検量線に照らし合わせて、調製直後の溶液Ａ１中のＦＧＦ－２３濃度ＣＡ１（調製直後）を決定した。調製直後の溶液Ａ１の代わりに、調製直後の溶液Ａ２～Ａ６の各溶液を用いて同様の測定を行い、調製直後の溶液Ａ２～Ａ６中のＦＧＦ－２３濃度ＣＡ２（調製直後）～ＣＡ６（調製直後）を決定した。また、調製直後の溶液Ａ１の代わりに、調製直後の溶液ａ０～ａ６の各溶液を用いて同様の測定を行い、調製直後の溶液ａ０～ａ６中のＦＧＦ－２３濃度Ｃａ０（調製直後）～Ｃａ６（調製直後）を決定した。 (6) Measurement of FGF-23 in each FGF-23 solution immediately after preparation (solutions A1 to A6; solutions a0 to a6) Solution A1 (0.5 mL) immediately after preparation in (1) above was added to (2) The sample for measurement was prepared by diluting 500 times with the antigen dilution solution prepared in (1). Using the prepared measurement sample, measurement was performed in the same manner as in (5) above, and the luminescence intensity for solution A1 was measured. The measured luminescence intensity was collated with the calibration curve prepared in (5) above to determine the FGF-23 concentration CA1 (immediately after preparation) in solution A1 immediately after preparation . The same measurement was performed using the solutions A2 to A6 immediately after preparation instead of the solution A1 immediately after preparation, and the FGF-23 concentration CA2 in the solutions A2 to A6 immediately after preparation CA2 (immediately after preparation) to CA6 (preparation ) Immediately after) . Further, instead of the solution A1 immediately after preparation, the same measurement was performed using the solutions a0 to a6 immediately after preparation, and the FGF-23 concentration Ca0 in the solutions a0 to a6 immediately after preparation Ca0 (immediately after preparation) to Ca6 (Immediately after preparation) was determined.
上記（１）で調製した直後の溶液Ａ１(0.5 mL)を、上記（２）で調製した抗原希釈液で500倍希釈し、測定用試料を調製した。調製した測定用試料を用いて、上記（５）と同様の方法により測定を行い、溶液Ａ１に対する発光強度を測定した。測定した発光強度を上記（５）で作成した検量線に照らし合わせて、調製直後の溶液Ａ１中のＦＧＦ－２３濃度ＣＡ１（調製直後）を決定した。調製直後の溶液Ａ１の代わりに、調製直後の溶液Ａ２～Ａ６の各溶液を用いて同様の測定を行い、調製直後の溶液Ａ２～Ａ６中のＦＧＦ－２３濃度ＣＡ２（調製直後）～ＣＡ６（調製直後）を決定した。また、調製直後の溶液Ａ１の代わりに、調製直後の溶液ａ０～ａ６の各溶液を用いて同様の測定を行い、調製直後の溶液ａ０～ａ６中のＦＧＦ－２３濃度Ｃａ０（調製直後）～Ｃａ６（調製直後）を決定した。 (6) Measurement of FGF-23 in each FGF-23 solution immediately after preparation (solutions A1 to A6; solutions a0 to a6) Solution A1 (0.5 mL) immediately after preparation in (1) above was added to (2) The sample for measurement was prepared by diluting 500 times with the antigen dilution solution prepared in (1). Using the prepared measurement sample, measurement was performed in the same manner as in (5) above, and the luminescence intensity for solution A1 was measured. The measured luminescence intensity was collated with the calibration curve prepared in (5) above to determine the FGF-23 concentration CA1 (immediately after preparation) in solution A1 immediately after preparation . The same measurement was performed using the solutions A2 to A6 immediately after preparation instead of the solution A1 immediately after preparation, and the FGF-23 concentration CA2 in the solutions A2 to A6 immediately after preparation CA2 (immediately after preparation) to CA6 (preparation ) Immediately after) . Further, instead of the solution A1 immediately after preparation, the same measurement was performed using the solutions a0 to a6 immediately after preparation, and the FGF-23 concentration Ca0 in the solutions a0 to a6 immediately after preparation Ca0 (immediately after preparation) to Ca6 (Immediately after preparation) was determined.
（７）4℃で7日間保存後の各ＦＧＦ－２３溶液（溶液Ａ１～Ａ６；溶液ａ０～ａ６）中のＦＧＦ－２３の測定
次に、上記（１）で調製したＦＧＦ－２３溶液（溶液Ａ１）(0.5 mL)をポリプロピレン製の容器内で、4℃で7日間保存した。調製直後の溶液Ａ１の代わりに、この4℃で7日間保存後のＦＧＦ－２３溶液Ａ１を用いて、上記（５）と同様の測定を行い、4℃で7日間保存後の溶液Ａ１中のＦＧＦ－２３濃度ＣＡ１（保存後）を決定した。同様に、4℃で7日間保存後の溶液Ａ２～Ａ６の各溶液を用いて測定を行い、4℃で7日間保存後の各溶液Ａ２～Ａ６中のＦＧＦ－２３濃度ＣＡ２（保存後）～ＣＡ６（保存後）を決定した。また、調製直後の溶液Ａ１の代わりに、4℃で7日間保存後の溶液ａ０～ａ６の各溶液を用いて同様の測定を行い、4℃で7日間保存後の溶液ａ０～ａ６中のＦＧＦ－２３濃度Ｃａ０（保存後）～Ｃａ６（保存後）を決定した。 (7) Measurement of FGF-23 in each FGF-23 solution (solutions A1 to A6; solutions a0 to a6) after storage at 4 ° C. for 7 days Next, the FGF-23 solution (solution) prepared in the above (1) A1) (0.5 mL) was stored in a polypropylene container at 4 ° C. for 7 days. Using the FGF-23 solution A1 stored at 4 ° C. for 7 days instead of the solution A1 immediately after preparation, the same measurement as in (5) above was performed, The FGF-23 concentration CA1 (after storage) was determined. Similarly, measurement was performed using each of solutions A2 to A6 after 7 days storage at 4 ° C., and FGF-23 concentration CA2 in each solution A2 to A6 after 7 days storage at 4 ° C. (after storage) CA6 (after storage) was determined. Further, instead of the solution A1 immediately after preparation, the same measurement was performed using the solutions a0 to a6 after storage at 4 ° C. for 7 days, and the FGF in the solutions a0 to a6 after storage at 4 ° C. for 7 days. -23 concentrations Ca0 (after storage) to Ca6 (after storage) were determined.
次に、上記（１）で調製したＦＧＦ－２３溶液（溶液Ａ１）(0.5 mL)をポリプロピレン製の容器内で、4℃で7日間保存した。調製直後の溶液Ａ１の代わりに、この4℃で7日間保存後のＦＧＦ－２３溶液Ａ１を用いて、上記（５）と同様の測定を行い、4℃で7日間保存後の溶液Ａ１中のＦＧＦ－２３濃度ＣＡ１（保存後）を決定した。同様に、4℃で7日間保存後の溶液Ａ２～Ａ６の各溶液を用いて測定を行い、4℃で7日間保存後の各溶液Ａ２～Ａ６中のＦＧＦ－２３濃度ＣＡ２（保存後）～ＣＡ６（保存後）を決定した。また、調製直後の溶液Ａ１の代わりに、4℃で7日間保存後の溶液ａ０～ａ６の各溶液を用いて同様の測定を行い、4℃で7日間保存後の溶液ａ０～ａ６中のＦＧＦ－２３濃度Ｃａ０（保存後）～Ｃａ６（保存後）を決定した。 (7) Measurement of FGF-23 in each FGF-23 solution (solutions A1 to A6; solutions a0 to a6) after storage at 4 ° C. for 7 days Next, the FGF-23 solution (solution) prepared in the above (1) A1) (0.5 mL) was stored in a polypropylene container at 4 ° C. for 7 days. Using the FGF-23 solution A1 stored at 4 ° C. for 7 days instead of the solution A1 immediately after preparation, the same measurement as in (5) above was performed, The FGF-23 concentration CA1 (after storage) was determined. Similarly, measurement was performed using each of solutions A2 to A6 after 7 days storage at 4 ° C., and FGF-23 concentration CA2 in each solution A2 to A6 after 7 days storage at 4 ° C. (after storage) CA6 (after storage) was determined. Further, instead of the solution A1 immediately after preparation, the same measurement was performed using the solutions a0 to a6 after storage at 4 ° C. for 7 days, and the FGF in the solutions a0 to a6 after storage at 4 ° C. for 7 days. -23 concentrations Ca0 (after storage) to Ca6 (after storage) were determined.
（８）保存後の各ＦＧＦ－２３溶液中のＦＧＦ－２３の残存率の決定
上記（６）で決定したＦＧＦ－２３濃度ＣＡｉ（調製直後）（ｉ：１～６）を100としたときの、上記（７）で決定したＦＧＦ－２３濃度ＣＡｉ（保存後）（ｉ：１～６）の相対値を第１表に示す。当該相対値は、4℃で7日間保存後のＦＧＦ－２３の残存率を表す。 (8) Determination of residual ratio of FGF-23 in each FGF-23 solution after storage When FGF-23 concentration CAi determined in (6) above ( immediately after preparation) (i: 1 to 6) is 100 The relative values of the FGF-23 concentration CAi (after storage) (i: 1 to 6) determined in (7) above are shown in Table 1. The relative value represents the residual ratio of FGF-23 after storage at 4 ° C. for 7 days.
上記（６）で決定したＦＧＦ－２３濃度ＣＡｉ（調製直後）（ｉ：１～６）を100としたときの、上記（７）で決定したＦＧＦ－２３濃度ＣＡｉ（保存後）（ｉ：１～６）の相対値を第１表に示す。当該相対値は、4℃で7日間保存後のＦＧＦ－２３の残存率を表す。 (8) Determination of residual ratio of FGF-23 in each FGF-23 solution after storage When FGF-23 concentration CAi determined in (6) above ( immediately after preparation) (i: 1 to 6) is 100 The relative values of the FGF-23 concentration CAi (after storage) (i: 1 to 6) determined in (7) above are shown in Table 1. The relative value represents the residual ratio of FGF-23 after storage at 4 ° C. for 7 days.
上記（６）で決定したＦＧＦ－２３濃度Ｃａｉ（調製直後）（ｉ：０～６）と、上記（７）で決定したＦＧＦ－２３濃度Ｃａｉ（保存後）（ｉ：０～６）とから、上記式（Ｉ）を用いて残存率（％）を算出した。その結果を第１表に示す。
FGF-23 concentration Cai determined in (6) above ( immediately after preparation) (i: 0 to 6) and FGF-23 concentration Cai determined in (7) above ( after storage) (i: 0 to 6) The residual ratio (%) was calculated using the above formula (I). The results are shown in Table 1.
残存率（％）が100に近い程、溶液中のＦＧＦ－２３が安定に保持されていることを示している。第１表の結果から明らかな様に、ポリオキシエチレン系の非イオン性界面活性剤を含有する溶液（溶液Ａ１～Ａ６）においては、いずれも残存率（％）が７０以上であるのに対して、ポリオキシエチレン系の非イオン性界面活性剤を含有しない溶液（溶液ａ０）、カチオン性界面活性剤を含有する溶液（溶液ａ１、ａ２）、陰イオン性界面活性剤を含有する溶液（溶液ａ３、ａ４）、及び、両性界面活性剤を含有する溶液（溶液ａ５、ａ６）においては、ＦＧＦ－２３の残存率は、0～5%であり、ＦＧＦ－２３は残存しないか、ほとんど残存しなかった。従って、ポリオキシエチレン系の非イオン性界面活性剤を含む本発明の安定化剤により、ＦＧＦ－２３が安定化されることが明らかとなった。
The closer the remaining rate (%) is to 100, the more stable the FGF-23 in the solution is. As is clear from the results in Table 1, in the solutions containing the polyoxyethylene-based nonionic surfactant (solutions A1 to A6), the residual ratio (%) is 70 or more in all cases. Solution containing no polyoxyethylene nonionic surfactant (solution a0), solution containing cationic surfactant (solution a1, a2), solution containing anionic surfactant (solution) In the solutions containing a3, a4) and amphoteric surfactants (solutions a5, a6), the residual ratio of FGF-23 is 0-5%, and FGF-23 does not remain or hardly remains. There wasn't. Therefore, it was revealed that FGF-23 is stabilized by the stabilizer of the present invention containing a polyoxyethylene-based nonionic surfactant.
ＢＳＡの添加によるＦＧＦ－２３の長期保存安定化
（１）ＦＧＦ－２３溶液（溶液Ａ７）の調製
以下の組成からなるＦＧＦ－２３溶液（溶液Ａ７）を調製した。
リン酸緩衝液 10 mmol/L (pH7.4)
ＦＧＦ－２３ 1μg/mL
塩化ナトリウム 140 mmol/L
Polysorbate 80 0.001% Stabilization of long-term storage of FGF-23 by addition of BSA (1) Preparation of FGF-23 solution (solution A7) An FGF-23 solution (solution A7) having the following composition was prepared.
Phosphate buffer 10 mmol / L (pH7.4)
FGF-23 1μg / mL
Sodium chloride 140 mmol / L
Polysorbate 80 0.001%
（１）ＦＧＦ－２３溶液（溶液Ａ７）の調製
以下の組成からなるＦＧＦ－２３溶液（溶液Ａ７）を調製した。
リン酸緩衝液 10 mmol/L (pH7.4)
ＦＧＦ－２３ 1μg/mL
塩化ナトリウム 140 mmol/L
Polysorbate 80 0.001% Stabilization of long-term storage of FGF-23 by addition of BSA (1) Preparation of FGF-23 solution (solution A7) An FGF-23 solution (solution A7) having the following composition was prepared.
Phosphate buffer 10 mmol / L (pH7.4)
FGF-23 1μg / mL
Sodium chloride 140 mmol / L
Polysorbate 80 0.001%
（２）ＦＧＦ－２３溶液（溶液Ｂ）の調製
以下の組成からなるＦＧＦ－２３溶液（溶液Ｂ）を調製した。
リン酸緩衝液 10 mmol/L (pH7.4)
ＦＧＦ－２３ 1μg/mL
塩化ナトリウム 140 mmol/L
Polysorbate 80 0.001%
ＢＳＡ 1% (2) Preparation of FGF-23 solution (solution B) An FGF-23 solution (solution B) having the following composition was prepared.
Phosphate buffer 10 mmol / L (pH7.4)
FGF-23 1μg / mL
Sodium chloride 140 mmol / L
Polysorbate 80 0.001%
BSA 1%
以下の組成からなるＦＧＦ－２３溶液（溶液Ｂ）を調製した。
リン酸緩衝液 10 mmol/L (pH7.4)
ＦＧＦ－２３ 1μg/mL
塩化ナトリウム 140 mmol/L
Polysorbate 80 0.001%
ＢＳＡ 1% (2) Preparation of FGF-23 solution (solution B) An FGF-23 solution (solution B) having the following composition was prepared.
Phosphate buffer 10 mmol / L (pH7.4)
FGF-23 1μg / mL
Sodium chloride 140 mmol / L
Polysorbate 80 0.001%
BSA 1%
（３）ＦＧＦ－２３溶液（溶液ａ０）の調製
以下の組成からなるＦＧＦ－２３溶液（溶液ａ０）を調製した。
リン酸緩衝液 10 mmol/L (pH7.4)
ＦＧＦ－２３ 1μg/mL
塩化ナトリウム 140 mmol/L (3) Preparation of FGF-23 solution (solution a0) An FGF-23 solution (solution a0) having the following composition was prepared.
Phosphate buffer 10 mmol / L (pH7.4)
FGF-23 1μg / mL
Sodium chloride 140 mmol / L
以下の組成からなるＦＧＦ－２３溶液（溶液ａ０）を調製した。
リン酸緩衝液 10 mmol/L (pH7.4)
ＦＧＦ－２３ 1μg/mL
塩化ナトリウム 140 mmol/L (3) Preparation of FGF-23 solution (solution a0) An FGF-23 solution (solution a0) having the following composition was prepared.
Phosphate buffer 10 mmol / L (pH7.4)
FGF-23 1μg / mL
Sodium chloride 140 mmol / L
（４）調製直後の各ＦＧＦ－２３溶液中のＦＧＦ－２３の測定
上記（１）で調製した直後のＦＧＦ－２３溶液（溶液Ａ７）中のＦＧＦ－２３濃度ＣＡ７（調製直後）を、実施例１の（４）に記載の方法により決定した。同様に、上記（２）で調製した直後のＦＧＦ－２３溶液（溶液Ｂ）中のＦＧＦ－２３濃度ＣＢ（調製直後）を決定した。さらに、上記（３）で調製した直後のＦＧＦ－２３溶液（溶液ａ０）中のＦＧＦ－２３濃度Ｃａ０（調製直後）を決定した。 (4) Measurement of FGF-23 in each FGF-23 solution immediately after preparation FGF-23 concentration CA7 (immediately after preparation ) in the FGF-23 solution (solution A7) immediately after preparation in (1) above It was determined by the method described in 1 (4). Similarly, the FGF-23 concentration CB (immediately after preparation ) in the FGF-23 solution (solution B) immediately after preparation in (2) above was determined. Furthermore, to determine the FGF-23 solution immediately after prepared in (3) (a solution a0) FGF-23 concentrations Ca0 (immediately after preparation) in.
上記（１）で調製した直後のＦＧＦ－２３溶液（溶液Ａ７）中のＦＧＦ－２３濃度ＣＡ７（調製直後）を、実施例１の（４）に記載の方法により決定した。同様に、上記（２）で調製した直後のＦＧＦ－２３溶液（溶液Ｂ）中のＦＧＦ－２３濃度ＣＢ（調製直後）を決定した。さらに、上記（３）で調製した直後のＦＧＦ－２３溶液（溶液ａ０）中のＦＧＦ－２３濃度Ｃａ０（調製直後）を決定した。 (4) Measurement of FGF-23 in each FGF-23 solution immediately after preparation FGF-23 concentration CA7 (immediately after preparation ) in the FGF-23 solution (solution A7) immediately after preparation in (1) above It was determined by the method described in 1 (4). Similarly, the FGF-23 concentration CB (immediately after preparation ) in the FGF-23 solution (solution B) immediately after preparation in (2) above was determined. Furthermore, to determine the FGF-23 solution immediately after prepared in (3) (a solution a0) FGF-23 concentrations Ca0 (immediately after preparation) in.
（５）40℃での保存後の各ＦＧＦ－２３溶液中のＦＧＦ－２３の測定
上記（１）で調製したＦＧＦ－２３溶液（溶液Ａ７）(0.5 mL)をポリプロピレン製の容器内で、40℃で保存した。40℃で1週間保存した後の溶液Ａ７中のＦＧＦ－２３濃度ＣＡ７（１週間保存後）を、実施例１の（５）に記載の方法により決定した。同様に、40℃で2週間保存した後の溶液Ａ７中のＦＧＦ－２３濃度ＣＡ７（２週間保存後）を決定した。ＦＧＦ－２３溶液（溶液Ａ７）の代わりに、上記（２）で調製したＦＧＦ－２３溶液（溶液Ｂ）を用いて、同様の測定を行い、40℃で1週間保存した後の溶液Ｂ中のＦＧＦ－２３濃度ＣＢ（１週間保存後）を、40℃で2週間保存した後の溶液Ｂ中のＦＧＦ－２３濃度ＣＢ（２週間保存後）を決定した。また、ＦＧＦ－２３溶液（溶液Ａ７）の代わりに、上記（３）で調製したＦＧＦ－２３溶液（溶液ａ０）を用いて、同様の測定を行い、40℃で1週間保存した後の溶液ａ０中のＦＧＦ－２３濃度Ｃａ０（１週間保存後）を、40℃で2週間保存した後の溶液ａ０中のＦＧＦ－２３濃度Ｃａ０（２週間保存後）を決定した。 (5) Measurement of FGF-23 in each FGF-23 solution after storage at 40 ° C. The FGF-23 solution (solution A7) (0.5 mL) prepared in (1) above was placed in a polypropylene container. Stored at ° C. The FGF-23 concentration CA7 (after storage for 1 week) in solution A7 after storage at 40 ° C. for 1 week was determined by the method described in Example 1, (5). Similarly, the FGF-23 concentration CA7 (after storage for 2 weeks) in solution A7 after storage at 40 ° C. for 2 weeks was determined. Using the FGF-23 solution (solution B) prepared in (2) above instead of the FGF-23 solution (solution A7), the same measurement was performed, and the solution B after storing at 40 ° C. for 1 week The FGF-23 concentration CB (after storage for 2 weeks ) in the solution B after 2 weeks storage at 40 ° C. was determined for the FGF-23 concentration CB (after storage for 1 week) . Further, instead of the FGF-23 solution (solution A7), the same measurement was performed using the FGF-23 solution (solution a0) prepared in (3) above, and the solution a0 after storage at 40 ° C. for 1 week was used. the FGF-23 concentrations Ca0 (after 1 week storage) in were determined 2 weeks saved FGF-23 concentration in the solution a0 after Ca0 (after storage for 2 weeks) at 40 ° C..
上記（１）で調製したＦＧＦ－２３溶液（溶液Ａ７）(0.5 mL)をポリプロピレン製の容器内で、40℃で保存した。40℃で1週間保存した後の溶液Ａ７中のＦＧＦ－２３濃度ＣＡ７（１週間保存後）を、実施例１の（５）に記載の方法により決定した。同様に、40℃で2週間保存した後の溶液Ａ７中のＦＧＦ－２３濃度ＣＡ７（２週間保存後）を決定した。ＦＧＦ－２３溶液（溶液Ａ７）の代わりに、上記（２）で調製したＦＧＦ－２３溶液（溶液Ｂ）を用いて、同様の測定を行い、40℃で1週間保存した後の溶液Ｂ中のＦＧＦ－２３濃度ＣＢ（１週間保存後）を、40℃で2週間保存した後の溶液Ｂ中のＦＧＦ－２３濃度ＣＢ（２週間保存後）を決定した。また、ＦＧＦ－２３溶液（溶液Ａ７）の代わりに、上記（３）で調製したＦＧＦ－２３溶液（溶液ａ０）を用いて、同様の測定を行い、40℃で1週間保存した後の溶液ａ０中のＦＧＦ－２３濃度Ｃａ０（１週間保存後）を、40℃で2週間保存した後の溶液ａ０中のＦＧＦ－２３濃度Ｃａ０（２週間保存後）を決定した。 (5) Measurement of FGF-23 in each FGF-23 solution after storage at 40 ° C. The FGF-23 solution (solution A7) (0.5 mL) prepared in (1) above was placed in a polypropylene container. Stored at ° C. The FGF-23 concentration CA7 (after storage for 1 week) in solution A7 after storage at 40 ° C. for 1 week was determined by the method described in Example 1, (5). Similarly, the FGF-23 concentration CA7 (after storage for 2 weeks) in solution A7 after storage at 40 ° C. for 2 weeks was determined. Using the FGF-23 solution (solution B) prepared in (2) above instead of the FGF-23 solution (solution A7), the same measurement was performed, and the solution B after storing at 40 ° C. for 1 week The FGF-23 concentration CB (after storage for 2 weeks ) in the solution B after 2 weeks storage at 40 ° C. was determined for the FGF-23 concentration CB (after storage for 1 week) . Further, instead of the FGF-23 solution (solution A7), the same measurement was performed using the FGF-23 solution (solution a0) prepared in (3) above, and the solution a0 after storage at 40 ° C. for 1 week was used. the FGF-23 concentrations Ca0 (after 1 week storage) in were determined 2 weeks saved FGF-23 concentration in the solution a0 after Ca0 (after storage for 2 weeks) at 40 ° C..
（６）40℃での保存後の各ＦＧＦ－２３溶液中のＦＧＦ－２３の残存率の決定
上記（４）で決定したＦＧＦ－２３濃度ＣＡ７（調製直後）と、上記（５）で決定したＦＧＦ－２３濃度ＣＡ７（１週間保存後）及びＣＡ７（２週間保存後）とから、式（Ｉ）により残存率（％）を算出した。その結果を第２表に示す。 (6) Determination of residual ratio of FGF-23 in each FGF-23 solution after storage at 40 ° C. FGF-23 concentration CA7 (immediately after preparation) determined in (4) above and determined in (5) above From the FGF-23 concentrations CA7 (after 1 week storage) and CA7 (after 2 weeks storage ), the residual rate (%) was calculated by the formula (I). The results are shown in Table 2.
上記（４）で決定したＦＧＦ－２３濃度ＣＡ７（調製直後）と、上記（５）で決定したＦＧＦ－２３濃度ＣＡ７（１週間保存後）及びＣＡ７（２週間保存後）とから、式（Ｉ）により残存率（％）を算出した。その結果を第２表に示す。 (6) Determination of residual ratio of FGF-23 in each FGF-23 solution after storage at 40 ° C. FGF-23 concentration CA7 (immediately after preparation) determined in (4) above and determined in (5) above From the FGF-23 concentrations CA7 (after 1 week storage) and CA7 (after 2 weeks storage ), the residual rate (%) was calculated by the formula (I). The results are shown in Table 2.
上記（４）で決定したＦＧＦ－２３濃度ＣＢ（調製直後）と、上記（５）で決定したＦＧＦ－２３濃度ＣＢ（１週間保存後）及びＣＢ（２週間保存後）とから、式（Ｉ）により残存率（％）を算出した。また、上記（４）で決定したＦＧＦ－２３濃度Ｃａ０（調製直後）と、上記（５）で決定したＦＧＦ－２３濃度Ｃａ０（１週間保存後）及びＣａ０（２週間保存後）とから、式（Ｉ）により残存率（％）を算出した。その結果を第２表に示す。
From the FGF-23 concentration CB determined in (4) above ( immediately after preparation) and the FGF-23 concentrations CB determined in (5) above ( after storage for 1 week) and CB (after storage for 2 weeks) , the formula (I ) To calculate the residual rate (%). Further, from the FGF-23 concentration Ca0 (immediately after preparation ) determined in (4) above, and the FGF-23 concentrations Ca0 (after storage for 1 week) and Ca0 (after storage for 2 weeks ) determined in (5) above, the formula The residual rate (%) was calculated from (I). The results are shown in Table 2.
第２表の結果から明らかな様に、ポリオキシエチレン系の非イオン性界面活性剤を含有しないＦＧＦ－２３溶液（溶液ａ０）においては、40℃で1週間保存後で、すでに、ＦＧＦ－２３が残存していないが、ポリオキシエチレン系の非イオン性界面活性剤と共にＢＳＡを含有するＦＧＦ－２３溶液（溶液Ｂ）においては、ポリオキシエチレン系の非イオン性界面活性剤を含有し、ＢＳＡを含有しないＦＧＦ－２３溶液（溶液Ａ７）に比較して、40℃で1週間保存後、及び、40℃で2週間保存後も、残存率が高かった。従って、本発明の、ポリオキシエチレン系の非イオン性界面活性剤及びＢＳＡを含有するＦＧＦ－２３の安定化剤を用いることにより、ＦＧＦ－２３がより安定化されることが明らかとなった。
As is apparent from the results in Table 2, in the FGF-23 solution (solution a0) containing no polyoxyethylene-based nonionic surfactant, FGF-23 was already stored after storage at 40 ° C. for 1 week. In the FGF-23 solution (solution B) containing BSA together with a polyoxyethylene nonionic surfactant, a polyoxyethylene nonionic surfactant is contained in the FGF-23 solution. Compared with the FGF-23 solution (solution A7) containing no lysate, the residual rate was high after storage at 40 ° C. for 1 week and also after storage at 40 ° C. for 2 weeks. Therefore, it has been clarified that FGF-23 is further stabilized by using the polyoxyethylene-based nonionic surfactant of the present invention and the FGF-23 stabilizer containing BSA.
本発明により、低リン血症性くる病、腫瘍性骨軟化症、腎不全等の疾患の診断に有効なＦＧＦ－２３の安定化剤、ＦＧＦ－２３の安定化方法、ＦＧＦ－２３の保存方法、及び、ＦＧＦ－２３測定用試薬が提供される。
INDUSTRIAL APPLICABILITY According to the present invention, an FGF-23 stabilizer effective for diagnosis of diseases such as hypophosphatemic rickets, neoplastic osteomalacia and renal failure, a method for stabilizing FGF-23, and a method for storing FGF-23 And a reagent for measuring FGF-23.
Claims (17)
- 有効成分としてポリオキシエチレン系の非イオン性界面活性剤を含有することを特徴とする、線維芽細胞増殖因子－２３の安定化剤。 A stabilizer for fibroblast growth factor-23, comprising a polyoxyethylene-based nonionic surfactant as an active ingredient.
- 前記ポリオキシエチレン系の非イオン性界面活性剤が、ポリオキシエチレンソルビタン脂肪酸エステル、ポリオキシエチレンアルキルエーテル、ポリオキシエチレンアルキルフェニルエーテル、及び、ポリオキシエチレンアルキルアミンからなる群より選ばれる、少なくとも１種の非イオン性界面活性剤である、請求項１記載の安定化剤。 The polyoxyethylene-based nonionic surfactant is selected from the group consisting of polyoxyethylene sorbitan fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene alkylphenyl ether, and polyoxyethylene alkylamine, at least 1 The stabilizer of claim 1, which is a species of nonionic surfactant.
- さらにアルブミンを含有する、請求項１又は２記載の安定化剤。 Furthermore, the stabilizer of Claim 1 or 2 containing albumin.
- 前記アルブミンがウシ血清アルブミンである、請求項３記載の安定化剤。 The stabilizer of claim 3, wherein the albumin is bovine serum albumin.
- 線維芽細胞増殖因子－２３を含有する試料を、容器中で、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体と共存させることを特徴とする、線維芽細胞増殖因子－２３の安定化方法。 A sample of fibroblast growth factor-23, characterized in that a sample containing fibroblast growth factor-23 coexists in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant. Stabilization method.
- 前記ポリオキシエチレン系の非イオン性界面活性剤が、ポリオキシエチレンソルビタン脂肪酸エステル、ポリオキシエチレンアルキルエーテル、ポリオキシエチレンアルキルフェニルエーテル、及び、ポリオキシエチレンアルキルアミンからなる群より選ばれる、少なくとも１種の非イオン性界面活性剤である、請求項５記載の安定化方法。 The polyoxyethylene-based nonionic surfactant is selected from the group consisting of polyoxyethylene sorbitan fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene alkylphenyl ether, and polyoxyethylene alkylamine, at least 1 The stabilization method according to claim 5, which is a kind of nonionic surfactant.
- 線維芽細胞増殖因子－２３を含有する試料を、さらにアルブミンと共存させる、請求項５又は６記載の安定化方法。 The stabilization method according to claim 5 or 6, wherein a sample containing fibroblast growth factor-23 is further allowed to coexist with albumin.
- 前記アルブミンがウシ血清アルブミンである、請求項７記載の安定化方法。 The stabilization method according to claim 7, wherein the albumin is bovine serum albumin.
- 前記容器がプラスチック製容器である、請求項５～８のいずれか一項に記載の安定化方法。 The stabilization method according to any one of claims 5 to 8, wherein the container is a plastic container.
- 前記プラスチック製容器が、ポリエチレン製容器、ポリプロピレン製容器、及び、ポリスチレン製容器からなる群より選ばれる容器である、請求項９記載の安定化方法。 The stabilization method according to claim 9, wherein the plastic container is a container selected from the group consisting of a polyethylene container, a polypropylene container, and a polystyrene container.
- 線維芽細胞増殖因子－２３を含有する試料を、容器中で、ポリオキシエチレン系の非イオン性界面活性剤を含有する水性媒体と共存させることを特徴とする、線維芽細胞増殖因子－２３の保存方法。 A sample of fibroblast growth factor-23, characterized in that a sample containing fibroblast growth factor-23 coexists in a container with an aqueous medium containing a polyoxyethylene-based nonionic surfactant. Preservation method.
- 前記ポリオキシエチレン系の非イオン性界面活性剤が、ポリオキシエチレンソルビタン脂肪酸エステル、ポリオキシエチレンアルキルエーテル、ポリオキシエチレンアルキルフェニルエーテル、及び、ポリオキシエチレンアルキルアミンからなる群より選ばれる、少なくとも１種の非イオン性界面活性剤である、請求項１１記載の保存方法。 The polyoxyethylene-based nonionic surfactant is selected from the group consisting of polyoxyethylene sorbitan fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene alkylphenyl ether, and polyoxyethylene alkylamine, at least 1 The storage method according to claim 11, which is a nonionic surfactant of a kind.
- 線維芽細胞増殖因子－２３を含有する試料を、さらにアルブミンと共存させる、請求項１１又は１２記載の保存方法。 The preservation method according to claim 11 or 12, wherein a sample containing fibroblast growth factor-23 is further allowed to coexist with albumin.
- 前記アルブミンがウシ血清アルブミンである、請求項１３記載の保存方法。 The storage method according to claim 13, wherein the albumin is bovine serum albumin.
- 前記容器が、プラスチック製容器である、請求項１１～１４のいずれか１項に記載の保存方法。 The storage method according to any one of claims 11 to 14, wherein the container is a plastic container.
- 前記プラスチック製容器が、ポリエチレン製容器、ポリプロピレン製容器、及び、ポリスチレン製容器からなる群より選ばれる何れかの容器である、請求項１５記載の保存方法。 The storage method according to claim 15, wherein the plastic container is any container selected from the group consisting of a polyethylene container, a polypropylene container, and a polystyrene container.
- 請求項１～４のいずれか一項に記載の線維芽細胞増殖因子－２３の安定化剤を含有することを特徴とする、線維芽細胞増殖因子－２３測定用試薬。 A reagent for measuring fibroblast growth factor-23, comprising the stabilizer for fibroblast growth factor-23 according to any one of claims 1 to 4.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2017556024A JP6741021B2 (en) | 2015-12-15 | 2016-12-09 | Fibroblast Growth Factor-23 Stabilizer |
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2015-243740 | 2015-12-15 | ||
| JP2015243740 | 2015-12-15 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| WO2017104567A1 true WO2017104567A1 (en) | 2017-06-22 |
Family
ID=59056574
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| PCT/JP2016/086710 WO2017104567A1 (en) | 2015-12-15 | 2016-12-09 | Agent for stabilizing fibroblast growth factor 23 |
Country Status (2)
| Country | Link |
|---|---|
| JP (1) | JP6741021B2 (en) |
| WO (1) | WO2017104567A1 (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2018221544A1 (en) * | 2017-05-31 | 2018-12-06 | 協和メデックス株式会社 | Measurement method for fibroblast growth factor–23, measurement reagent, and measurement kit |
| WO2022215341A1 (en) * | 2021-04-05 | 2022-10-13 | 株式会社島津製作所 | METHOD FOR CREATING CALIBRATION CURVE AND METHOD FOR MEASURING AMYLOID β-RELATED PEPTIDE |
| JP7502169B2 (en) | 2020-12-11 | 2024-06-18 | アークレイ株式会社 | Hemoglobin measurement reagents |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH02138223A (en) * | 1988-06-06 | 1990-05-28 | Takeda Chem Ind Ltd | Stabilized composition of fibroblast growth factor or mucin thereof and preparation thereof |
| JPH0782171A (en) * | 1993-09-17 | 1995-03-28 | Teikoku Seiyaku Co Ltd | Film preparation containing fibroblast growth factor |
| JPH11158076A (en) * | 1997-12-01 | 1999-06-15 | Asahi Medical Co Ltd | Specific removing material for cell |
| JP2009234988A (en) * | 2008-03-27 | 2009-10-15 | Sanyo Chem Ind Ltd | Cardiac muscle troponin t-containing aqueous solution and production method thereof |
-
2016
- 2016-12-09 WO PCT/JP2016/086710 patent/WO2017104567A1/en active Application Filing
- 2016-12-09 JP JP2017556024A patent/JP6741021B2/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH02138223A (en) * | 1988-06-06 | 1990-05-28 | Takeda Chem Ind Ltd | Stabilized composition of fibroblast growth factor or mucin thereof and preparation thereof |
| JPH0782171A (en) * | 1993-09-17 | 1995-03-28 | Teikoku Seiyaku Co Ltd | Film preparation containing fibroblast growth factor |
| JPH11158076A (en) * | 1997-12-01 | 1999-06-15 | Asahi Medical Co Ltd | Specific removing material for cell |
| JP2009234988A (en) * | 2008-03-27 | 2009-10-15 | Sanyo Chem Ind Ltd | Cardiac muscle troponin t-containing aqueous solution and production method thereof |
Non-Patent Citations (1)
| Title |
|---|
| ITOH NOBUYUKI ET AL., EVOLUTION OF THE FGF AND FGFR GENE FAMILIES, TRENDS IN GENETICS, vol. 20, no. 11, 2004, pages 563 - 569, XP004593427 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2018221544A1 (en) * | 2017-05-31 | 2018-12-06 | 協和メデックス株式会社 | Measurement method for fibroblast growth factor–23, measurement reagent, and measurement kit |
| JP7502169B2 (en) | 2020-12-11 | 2024-06-18 | アークレイ株式会社 | Hemoglobin measurement reagents |
| WO2022215341A1 (en) * | 2021-04-05 | 2022-10-13 | 株式会社島津製作所 | METHOD FOR CREATING CALIBRATION CURVE AND METHOD FOR MEASURING AMYLOID β-RELATED PEPTIDE |
| JP7563580B2 (en) | 2021-04-05 | 2024-10-08 | 株式会社島津製作所 | Method for creating a calibration curve and method for measuring amyloid-β-related peptide |
Also Published As
| Publication number | Publication date |
|---|---|
| JPWO2017104567A1 (en) | 2018-10-04 |
| JP6741021B2 (en) | 2020-08-19 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| JP7541499B2 (en) | Virus inactivation method using environmentally friendly cleaning agents | |
| JP6014091B2 (en) | Method for measuring component to be measured in sample and measurement kit | |
| JP6741021B2 (en) | Fibroblast Growth Factor-23 Stabilizer | |
| US9663816B2 (en) | Method for measuring a component of a biological fluid and reducing the effect of interfering substances | |
| JP5653216B2 (en) | Cystatin C adsorption inhibitor | |
| JP4704355B2 (en) | Standard product for fractional determination of components in lipoproteins | |
| CA2872006C (en) | Method for stabilizing ascorbic acid oxidase | |
| JP5832723B2 (en) | Standard for quantification of soluble interleukin-2 receptor | |
| JP7084589B2 (en) | Oxidized high-density lipoprotein measurement method | |
| WO2016159203A1 (en) | Preservative solution for heme protein, and method for stabilizing heme protein | |
| WO2024204714A1 (en) | Reagent, kit, and method for detecting substance to be measured | |
| JP2018155629A (en) | Peptide adsorption inhibitor | |
| WO2018221544A1 (en) | Measurement method for fibroblast growth factor–23, measurement reagent, and measurement kit | |
| JP2014039486A (en) | Luminous enhancing method using n-(2-acetamide) iminodiacetic acid | |
| WO2022154121A1 (en) | Adsorption inhibitor for hepcidin, method for inhibiting adsorption, reference standard, reagent, kit and measurement method | |
| JP5326331B2 (en) | Method for stabilizing tyramine oxidase and composition thereof | |
| JP2009072105A (en) | Cholesterol-measuring reagent, enzyme-containing reagent, enzyme stabilizer and method for stabilizing enzyme | |
| JP2024046375A (en) | Antibody solution capable of suppressing non-specific reactions | |
| JP2024093998A (en) | Antibody solution capable of suppressing non-specific reaction | |
| JP2018169273A (en) | Peptide adsorption inhibitor | |
| JP2009240289A (en) | Method for stabilizing tyramine oxidase and composition thereof | |
| WO2014034822A1 (en) | Method for stabilizing catalase | |
| JP2009247256A (en) | Method for stabilizing tyramine oxidase and composition thereof | |
| JP2009247258A (en) | Method for stabilizing tyramine oxidase and composition thereof | |
| JP2010148526A (en) | Quantative method for cholesterol in low density lipoprotein |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| 121 | Ep: the epo has been informed by wipo that ep was designated in this application |
Ref document number: 16875547 Country of ref document: EP Kind code of ref document: A1 |
|
| ENP | Entry into the national phase |
Ref document number: 2017556024 Country of ref document: JP Kind code of ref document: A |
|
| NENP | Non-entry into the national phase |
Ref country code: DE |
|
| 122 | Ep: pct application non-entry in european phase |
Ref document number: 16875547 Country of ref document: EP Kind code of ref document: A1 |