JPH02206763A - Analysis for selective measurement of glycolipid composite in blood having sialic acid - Google Patents
Analysis for selective measurement of glycolipid composite in blood having sialic acidInfo
- Publication number
- JPH02206763A JPH02206763A JP1131718A JP13171889A JPH02206763A JP H02206763 A JPH02206763 A JP H02206763A JP 1131718 A JP1131718 A JP 1131718A JP 13171889 A JP13171889 A JP 13171889A JP H02206763 A JPH02206763 A JP H02206763A
- Authority
- JP
- Japan
- Prior art keywords
- precipitate
- sialic acid
- solution
- methanol
- water
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- SQVRNKJHWKZAKO-UHFFFAOYSA-N beta-N-Acetyl-D-neuraminic acid Natural products CC(=O)NC1C(O)CC(O)(C(O)=O)OC1C(O)C(O)CO SQVRNKJHWKZAKO-UHFFFAOYSA-N 0.000 title claims abstract description 132
- SQVRNKJHWKZAKO-OQPLDHBCSA-N sialic acid Chemical compound CC(=O)N[C@@H]1[C@@H](O)C[C@@](O)(C(O)=O)OC1[C@H](O)[C@H](O)CO SQVRNKJHWKZAKO-OQPLDHBCSA-N 0.000 title claims abstract description 129
- 210000004369 blood Anatomy 0.000 title claims abstract description 17
- 239000008280 blood Substances 0.000 title claims abstract description 17
- 229930186217 Glycolipid Natural products 0.000 title claims abstract description 15
- HVCOBJNICQPDBP-UHFFFAOYSA-N 3-[3-[3,5-dihydroxy-6-methyl-4-(3,4,5-trihydroxy-6-methyloxan-2-yl)oxyoxan-2-yl]oxydecanoyloxy]decanoic acid;hydrate Chemical compound O.OC1C(OC(CC(=O)OC(CCCCCCC)CC(O)=O)CCCCCCC)OC(C)C(O)C1OC1C(O)C(O)C(O)C(C)O1 HVCOBJNICQPDBP-UHFFFAOYSA-N 0.000 title claims abstract description 10
- 238000004458 analytical method Methods 0.000 title claims description 16
- 238000005259 measurement Methods 0.000 title description 9
- 239000002131 composite material Substances 0.000 title 1
- 210000002966 serum Anatomy 0.000 claims abstract description 92
- 238000000034 method Methods 0.000 claims abstract description 23
- 150000003839 salts Chemical class 0.000 claims abstract description 19
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 11
- 229910052751 metal Inorganic materials 0.000 claims abstract description 9
- 239000002184 metal Substances 0.000 claims abstract description 9
- 150000002632 lipids Chemical class 0.000 claims abstract description 8
- 229910021645 metal ion Inorganic materials 0.000 claims abstract description 7
- 230000007935 neutral effect Effects 0.000 claims abstract description 7
- 102000004895 Lipoproteins Human genes 0.000 claims abstract 2
- 108090001030 Lipoproteins Proteins 0.000 claims abstract 2
- 239000002244 precipitate Substances 0.000 claims description 183
- 239000000243 solution Substances 0.000 claims description 182
- 238000000605 extraction Methods 0.000 claims description 24
- 239000011259 mixed solution Substances 0.000 claims description 4
- 239000003791 organic solvent mixture Substances 0.000 claims description 4
- BHPQYMZQTOCNFJ-UHFFFAOYSA-N Calcium cation Chemical compound [Ca+2] BHPQYMZQTOCNFJ-UHFFFAOYSA-N 0.000 claims description 2
- JPVYNHNXODAKFH-UHFFFAOYSA-N Cu2+ Chemical compound [Cu+2] JPVYNHNXODAKFH-UHFFFAOYSA-N 0.000 claims description 2
- CWYNVVGOOAEACU-UHFFFAOYSA-N Fe2+ Chemical compound [Fe+2] CWYNVVGOOAEACU-UHFFFAOYSA-N 0.000 claims description 2
- VTLYFUHAOXGGBS-UHFFFAOYSA-N Fe3+ Chemical compound [Fe+3] VTLYFUHAOXGGBS-UHFFFAOYSA-N 0.000 claims description 2
- JLVVSXFLKOJNIY-UHFFFAOYSA-N Magnesium ion Chemical compound [Mg+2] JLVVSXFLKOJNIY-UHFFFAOYSA-N 0.000 claims description 2
- FOIXSVOLVBLSDH-UHFFFAOYSA-N Silver ion Chemical compound [Ag+] FOIXSVOLVBLSDH-UHFFFAOYSA-N 0.000 claims description 2
- PTFCDOFLOPIGGS-UHFFFAOYSA-N Zinc dication Chemical compound [Zn+2] PTFCDOFLOPIGGS-UHFFFAOYSA-N 0.000 claims description 2
- 229910052783 alkali metal Inorganic materials 0.000 claims description 2
- 150000001340 alkali metals Chemical class 0.000 claims description 2
- 239000008346 aqueous phase Substances 0.000 claims description 2
- WLZRMCYVCSSEQC-UHFFFAOYSA-N cadmium(2+) Chemical compound [Cd+2] WLZRMCYVCSSEQC-UHFFFAOYSA-N 0.000 claims description 2
- 239000011575 calcium Substances 0.000 claims description 2
- 229910001424 calcium ion Inorganic materials 0.000 claims description 2
- 229910001429 cobalt ion Inorganic materials 0.000 claims description 2
- XLJKHNWPARRRJB-UHFFFAOYSA-N cobalt(2+) Chemical compound [Co+2] XLJKHNWPARRRJB-UHFFFAOYSA-N 0.000 claims description 2
- 229910001447 ferric ion Inorganic materials 0.000 claims description 2
- 229910001448 ferrous ion Inorganic materials 0.000 claims description 2
- 229910001425 magnesium ion Inorganic materials 0.000 claims description 2
- 229910001453 nickel ion Inorganic materials 0.000 claims description 2
- 230000001376 precipitating effect Effects 0.000 claims description 2
- VEQPNABPJHWNSG-UHFFFAOYSA-N Nickel(2+) Chemical compound [Ni+2] VEQPNABPJHWNSG-UHFFFAOYSA-N 0.000 claims 1
- BQPIGGFYSBELGY-UHFFFAOYSA-N mercury(2+) Chemical compound [Hg+2] BQPIGGFYSBELGY-UHFFFAOYSA-N 0.000 claims 1
- 206010028980 Neoplasm Diseases 0.000 abstract description 17
- IYDGMDWEHDFVQI-UHFFFAOYSA-N phosphoric acid;trioxotungsten Chemical compound O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.O=[W](=O)=O.OP(O)(O)=O IYDGMDWEHDFVQI-UHFFFAOYSA-N 0.000 abstract description 3
- 238000001556 precipitation Methods 0.000 abstract description 3
- 239000002253 acid Substances 0.000 abstract 2
- 239000007788 liquid Substances 0.000 abstract 2
- 239000011877 solvent mixture Substances 0.000 abstract 2
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 516
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 174
- 239000000203 mixture Substances 0.000 description 97
- 239000006228 supernatant Substances 0.000 description 83
- 208000014829 head and neck neoplasm Diseases 0.000 description 20
- 239000000523 sample Substances 0.000 description 20
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L Magnesium chloride Chemical compound [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 description 16
- YKYOUMDCQGMQQO-UHFFFAOYSA-L cadmium dichloride Chemical compound Cl[Cd]Cl YKYOUMDCQGMQQO-UHFFFAOYSA-L 0.000 description 16
- 238000002156 mixing Methods 0.000 description 15
- 238000012360 testing method Methods 0.000 description 15
- 206010006187 Breast cancer Diseases 0.000 description 13
- 208000026310 Breast neoplasm Diseases 0.000 description 13
- 206010061535 Ovarian neoplasm Diseases 0.000 description 12
- 208000029742 colonic neoplasm Diseases 0.000 description 12
- JIAARYAFYJHUJI-UHFFFAOYSA-L zinc dichloride Chemical compound [Cl-].[Cl-].[Zn+2] JIAARYAFYJHUJI-UHFFFAOYSA-L 0.000 description 12
- 206010009944 Colon cancer Diseases 0.000 description 11
- 206010033128 Ovarian cancer Diseases 0.000 description 11
- 239000012154 double-distilled water Substances 0.000 description 11
- GHMLBKRAJCXXBS-UHFFFAOYSA-N resorcinol Chemical compound OC1=CC=CC(O)=C1 GHMLBKRAJCXXBS-UHFFFAOYSA-N 0.000 description 11
- ARUVKPQLZAKDPS-UHFFFAOYSA-L copper(II) sulfate Chemical compound [Cu+2].[O-][S+2]([O-])([O-])[O-] ARUVKPQLZAKDPS-UHFFFAOYSA-L 0.000 description 10
- 229910000366 copper(II) sulfate Inorganic materials 0.000 description 10
- 208000026037 malignant tumor of neck Diseases 0.000 description 10
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 9
- 208000009956 adenocarcinoma Diseases 0.000 description 9
- 239000001110 calcium chloride Substances 0.000 description 9
- 229910001628 calcium chloride Inorganic materials 0.000 description 9
- 238000002835 absorbance Methods 0.000 description 8
- 239000011790 ferrous sulphate Substances 0.000 description 8
- 235000003891 ferrous sulphate Nutrition 0.000 description 8
- BAUYGSIQEAFULO-UHFFFAOYSA-L iron(2+) sulfate (anhydrous) Chemical compound [Fe+2].[O-]S([O-])(=O)=O BAUYGSIQEAFULO-UHFFFAOYSA-L 0.000 description 8
- 229910000359 iron(II) sulfate Inorganic materials 0.000 description 8
- 229910001629 magnesium chloride Inorganic materials 0.000 description 8
- 229960002523 mercuric chloride Drugs 0.000 description 8
- LWJROJCJINYWOX-UHFFFAOYSA-L mercury dichloride Chemical compound Cl[Hg]Cl LWJROJCJINYWOX-UHFFFAOYSA-L 0.000 description 8
- 229910021586 Nickel(II) chloride Inorganic materials 0.000 description 7
- 201000010099 disease Diseases 0.000 description 7
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 7
- RLJMLMKIBZAXJO-UHFFFAOYSA-N lead nitrate Chemical compound [O-][N+](=O)O[Pb]O[N+]([O-])=O RLJMLMKIBZAXJO-UHFFFAOYSA-N 0.000 description 7
- QMMRZOWCJAIUJA-UHFFFAOYSA-L nickel dichloride Chemical compound Cl[Ni]Cl QMMRZOWCJAIUJA-UHFFFAOYSA-L 0.000 description 7
- 150000002339 glycosphingolipids Chemical class 0.000 description 6
- 229960001755 resorcinol Drugs 0.000 description 6
- 239000011592 zinc chloride Substances 0.000 description 6
- 235000005074 zinc chloride Nutrition 0.000 description 6
- NWONKYPBYAMBJT-UHFFFAOYSA-L zinc sulfate Chemical compound [Zn+2].[O-]S([O-])(=O)=O NWONKYPBYAMBJT-UHFFFAOYSA-L 0.000 description 5
- 229960001763 zinc sulfate Drugs 0.000 description 5
- 229910000368 zinc sulfate Inorganic materials 0.000 description 5
- 201000011510 cancer Diseases 0.000 description 4
- 238000006243 chemical reaction Methods 0.000 description 4
- SQGYOTSLMSWVJD-UHFFFAOYSA-N silver(1+) nitrate Chemical compound [Ag+].[O-]N(=O)=O SQGYOTSLMSWVJD-UHFFFAOYSA-N 0.000 description 4
- 238000003756 stirring Methods 0.000 description 4
- 206010000234 Abortion spontaneous Diseases 0.000 description 3
- 239000007864 aqueous solution Substances 0.000 description 3
- 208000015994 miscarriage Diseases 0.000 description 3
- 125000005629 sialic acid group Chemical group 0.000 description 3
- 208000000995 spontaneous abortion Diseases 0.000 description 3
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 3
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- 206010062717 Increased upper airway secretion Diseases 0.000 description 2
- 206010061218 Inflammation Diseases 0.000 description 2
- LRHPLDYGYMQRHN-UHFFFAOYSA-N N-Butanol Chemical compound CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 description 2
- PXHVJJICTQNCMI-UHFFFAOYSA-N Nickel Chemical compound [Ni] PXHVJJICTQNCMI-UHFFFAOYSA-N 0.000 description 2
- LCTONWCANYUPML-UHFFFAOYSA-M Pyruvate Chemical compound CC(=O)C([O-])=O LCTONWCANYUPML-UHFFFAOYSA-M 0.000 description 2
- 239000007983 Tris buffer Substances 0.000 description 2
- 230000002159 abnormal effect Effects 0.000 description 2
- 206010000210 abortion Diseases 0.000 description 2
- 231100000176 abortion Toxicity 0.000 description 2
- 206010003246 arthritis Diseases 0.000 description 2
- 238000004364 calculation method Methods 0.000 description 2
- 238000005119 centrifugation Methods 0.000 description 2
- 229940106189 ceramide Drugs 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 238000003745 diagnosis Methods 0.000 description 2
- 238000010790 dilution Methods 0.000 description 2
- 239000012895 dilution Substances 0.000 description 2
- 150000002270 gangliosides Chemical class 0.000 description 2
- 230000004054 inflammatory process Effects 0.000 description 2
- 230000008606 intracellular interaction Effects 0.000 description 2
- 239000012528 membrane Substances 0.000 description 2
- 230000001613 neoplastic effect Effects 0.000 description 2
- 150000002482 oligosaccharides Polymers 0.000 description 2
- 239000012071 phase Substances 0.000 description 2
- 208000026435 phlegm Diseases 0.000 description 2
- 238000005375 photometry Methods 0.000 description 2
- 229910001961 silver nitrate Inorganic materials 0.000 description 2
- 239000012086 standard solution Substances 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- YDNKGFDKKRUKPY-JHOUSYSJSA-N C16 ceramide Natural products CCCCCCCCCCCCCCCC(=O)N[C@@H](CO)[C@H](O)C=CCCCCCCCCCCCCC YDNKGFDKKRUKPY-JHOUSYSJSA-N 0.000 description 1
- 201000009030 Carcinoma Diseases 0.000 description 1
- 206010007882 Cellulitis Diseases 0.000 description 1
- 206010008342 Cervix carcinoma Diseases 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000001390 Fructose-Bisphosphate Aldolase Human genes 0.000 description 1
- 108010068561 Fructose-Bisphosphate Aldolase Proteins 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 102000003886 Glycoproteins Human genes 0.000 description 1
- 108090000288 Glycoproteins Proteins 0.000 description 1
- 102100038736 Histone H3.3C Human genes 0.000 description 1
- 101001031505 Homo sapiens Histone H3.3C Proteins 0.000 description 1
- 206010027476 Metastases Diseases 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- OVRNDRQMDRJTHS-CBQIKETKSA-N N-Acetyl-D-Galactosamine Chemical compound CC(=O)N[C@H]1[C@@H](O)O[C@H](CO)[C@H](O)[C@@H]1O OVRNDRQMDRJTHS-CBQIKETKSA-N 0.000 description 1
- MBLBDJOUHNCFQT-UHFFFAOYSA-N N-acetyl-D-galactosamine Natural products CC(=O)NC(C=O)C(O)C(O)C(O)CO MBLBDJOUHNCFQT-UHFFFAOYSA-N 0.000 description 1
- SQVRNKJHWKZAKO-PFQGKNLYSA-N N-acetyl-beta-neuraminic acid Chemical compound CC(=O)N[C@@H]1[C@@H](O)C[C@@](O)(C(O)=O)O[C@H]1[C@H](O)[C@H](O)CO SQVRNKJHWKZAKO-PFQGKNLYSA-N 0.000 description 1
- 206010061309 Neoplasm progression Diseases 0.000 description 1
- 102000005348 Neuraminidase Human genes 0.000 description 1
- 108010006232 Neuraminidase Proteins 0.000 description 1
- 206010033799 Paralysis Diseases 0.000 description 1
- 102000003992 Peroxidases Human genes 0.000 description 1
- 201000004681 Psoriasis Diseases 0.000 description 1
- 108010042687 Pyruvate Oxidase Proteins 0.000 description 1
- 208000033781 Thyroid carcinoma Diseases 0.000 description 1
- 208000024770 Thyroid neoplasm Diseases 0.000 description 1
- 208000025865 Ulcer Diseases 0.000 description 1
- 208000006105 Uterine Cervical Neoplasms Diseases 0.000 description 1
- 239000004015 abortifacient agent Substances 0.000 description 1
- 231100000641 abortifacient agent Toxicity 0.000 description 1
- KBGAYAKRZNYFFG-BOHATCBPSA-N aceneuramic acid Chemical compound OC(=O)C(=O)C[C@H](O)[C@@H](NC(=O)C)[C@@H](O)[C@H](O)[C@H](O)CO KBGAYAKRZNYFFG-BOHATCBPSA-N 0.000 description 1
- WQZGKKKJIJFFOK-PHYPRBDBSA-N alpha-D-galactose Chemical compound OC[C@H]1O[C@H](O)[C@H](O)[C@@H](O)[C@H]1O WQZGKKKJIJFFOK-PHYPRBDBSA-N 0.000 description 1
- 125000003277 amino group Chemical group 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
- 150000001661 cadmium Chemical class 0.000 description 1
- 238000011088 calibration curve Methods 0.000 description 1
- 239000002775 capsule Substances 0.000 description 1
- ZVEQCJWYRWKARO-UHFFFAOYSA-N ceramide Natural products CCCCCCCCCCCCCCC(O)C(=O)NC(CO)C(O)C=CCCC=C(C)CCCCCCCCC ZVEQCJWYRWKARO-UHFFFAOYSA-N 0.000 description 1
- 125000001549 ceramide group Chemical group 0.000 description 1
- 150000001783 ceramides Chemical class 0.000 description 1
- 201000010881 cervical cancer Diseases 0.000 description 1
- MXOAEAUPQDYUQM-UHFFFAOYSA-N chlorphenesin Chemical group OCC(O)COC1=CC=C(Cl)C=C1 MXOAEAUPQDYUQM-UHFFFAOYSA-N 0.000 description 1
- 239000013068 control sample Substances 0.000 description 1
- 238000012937 correction Methods 0.000 description 1
- 230000007423 decrease Effects 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000006911 enzymatic reaction Methods 0.000 description 1
- 229930182830 galactose Natural products 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 230000003862 health status Effects 0.000 description 1
- FUZZWVXGSFPDMH-UHFFFAOYSA-N hexanoic acid Chemical compound CCCCCC(O)=O FUZZWVXGSFPDMH-UHFFFAOYSA-N 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 229920002521 macromolecule Polymers 0.000 description 1
- 239000011777 magnesium Substances 0.000 description 1
- 230000003211 malignant effect Effects 0.000 description 1
- 150000007522 mineralic acids Chemical class 0.000 description 1
- 208000025440 neoplasm of neck Diseases 0.000 description 1
- VVGIYYKRAMHVLU-UHFFFAOYSA-N newbouldiamide Natural products CCCCCCCCCCCCCCCCCCCC(O)C(O)C(O)C(CO)NC(=O)CCCCCCCCCCCCCCCCC VVGIYYKRAMHVLU-UHFFFAOYSA-N 0.000 description 1
- 229920001542 oligosaccharide Polymers 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 230000001314 paroxysmal effect Effects 0.000 description 1
- 230000001575 pathological effect Effects 0.000 description 1
- 108040007629 peroxidase activity proteins Proteins 0.000 description 1
- 150000002978 peroxides Chemical class 0.000 description 1
- 235000015047 pilsener Nutrition 0.000 description 1
- 238000005498 polishing Methods 0.000 description 1
- 230000035935 pregnancy Effects 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 229910052709 silver Inorganic materials 0.000 description 1
- 239000004332 silver Substances 0.000 description 1
- -1 silver ions Chemical class 0.000 description 1
- 150000003408 sphingolipids Chemical class 0.000 description 1
- 239000011550 stock solution Substances 0.000 description 1
- 239000004575 stone Substances 0.000 description 1
- 239000013595 supernatant sample Substances 0.000 description 1
- 201000002510 thyroid cancer Diseases 0.000 description 1
- 208000013077 thyroid gland carcinoma Diseases 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 230000005751 tumor progression Effects 0.000 description 1
- 231100000397 ulcer Toxicity 0.000 description 1
- 235000021122 unsaturated fatty acids Nutrition 0.000 description 1
- 150000004670 unsaturated fatty acids Chemical class 0.000 description 1
- 239000011701 zinc Substances 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
- G01N33/52—Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper and including single- and multilayer analytical elements
-
- 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
- G01N33/92—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving lipids, e.g. cholesterol, lipoproteins, or their receptors
-
- 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
- G01N33/66—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving blood sugars, e.g. galactose
Abstract
Description
【発明の詳細な説明】
〔産業上の利用分野〕
本発明は、シアル酸を有する糖脂質複合体の選択的な測
定を目的とする分析方法に関する。DETAILED DESCRIPTION OF THE INVENTION [Industrial Application Field] The present invention relates to an analytical method for selectively measuring a glycolipid complex having sialic acid.
シアル酸（Ｎ−アセチルノイラミン酸）は、生体内では
、遊離状態でも、結合状態でも存在するが［アドバンセ
ズ・イン・キャンサー・リサーチ（Ａｄｖａｎ。Sialic acid (N-acetylneuraminic acid) exists in both free and bound states in living organisms [Advances in Cancer Research (Advances).
Ｃａｎｃｅｒ Ｒｅｓ、）、３８巻（１９８３年）、２
８９〜３５０ベージ］、特に、オリゴ糖鎖の末端基とな
ることにより、これを介して、膜結合性スフィンゴ脂質
と結合することがある［ジャーナル・オブ・スープラモ
レキュラー・ストラフチャー（Ｊ、５ｕｐｒａ＋５ｏｌ
ｓｃｕｌａｒ ５ｔｒｕｃｔｕｒｅ）。Cancer Res, ), vol. 38 (1983), 2
89-350 pages], and in particular may bind to membrane-bound sphingolipids by forming terminal groups of oligosaccharide chains [Journal of Supramolecular Stretchers (J, 5upra+5ol
5 structures).
９巻（１９７８年）、 １５７〜１７７ページコ。Volume 9 (1978), pages 157-177.
スフィンゴ糖脂質も含めたガングリオシドなる用語は１
文献中において必ずしも明確なわけではない、スベンネ
ルホルム（Ｓｖａｎｎｓｒｈｏｌａ＋） ［コンプリヘ
ンシブ０バイオケミストリー（Ｃｏｍｐｒｅｈｅｎｓｉ
ｖｅＢｉｏｃｈｅｍ、）、 １８巻（１９７０年）、２
０１ページ、エルゼビア社（アムステルダム）発行］に
よれば、このように呼ばれる化合物は、モノシアロガン
グリオシド、ジシアロガングリオシド、および、トリジ
アロガングリオシドなる群に分類される。ガングリオシ
ドの疎水性の部分はセラミドとよばれ、その不飽和脂肪
酸の長い分子鎖によって膜と結合している。The term ganglioside, which also includes glycosphingolipids, is 1
Svannsrhola+ [Comprehensive Biochemistry] is not always clear in the literature.
veBiochem, ), Volume 18 (1970), 2
01, published by Elsevier (Amsterdam)], the compounds so called are classified into the groups monosialogangliosides, disialogangliosides, and tridialogangliosides. The hydrophobic part of gangliosides, called ceramides, is bound to membranes by long chains of unsaturated fatty acids.
セラミドは、アミノ基を介して、オリゴ糖類、すなわち
、グルコース、ガラクトース、Ｎ−アセチルガラクトサ
ミン、シアル酸などと結合している。Ceramide is bonded to oligosaccharides, ie, glucose, galactose, N-acetylgalactosamine, sialic acid, etc., via an amino group.
スフィンゴ糖脂質の巨大分子は、これにシアル酸が結合
しているため、負に荷電しており、この負電荷が、ある
種の細胞内相互作用、に影響を及ぼす。Glycosphingolipid macromolecules are negatively charged due to the binding of sialic acid to them, and this negative charge affects certain intracellular interactions.
腫瘍が存在する器官においては、生化学的平衡の乱れか
ら細胞内相互作用が変化し、そのため、糖脂質に結合し
たシアル酸の血液中の量が増大する。In organs where tumors exist, intracellular interactions change due to disturbances in the biochemical equilibrium, resulting in an increase in the amount of sialic acid bound to glycolipids in the blood.
上記の理論的認識に基づき、糖脂質と結合したシアル酸
の血中含有量の測定を目的とする方法が、いくつか案出
されている。Based on the above theoretical understanding, several methods have been devised for the purpose of measuring the blood content of sialic acid bound to glycolipids.
スベンネルホルムの方法［ビオケミ力・エト・ビオフィ
ジ力・アクタ（Ｂｉｏｃｈｅｔｍ、Ｂｉｏｐｈｙｓ、Ａ
ｃｔａ）、 ２４巻（１９５７年）、６０４〜６１１ペ
ージ］によれば、クロロホルムとメタノールの２：１混
合液を用い、血漿に対する抽出を行なって、スフィンゴ
糖脂質を、血中の他の脂質分画から分離し、第二銅化合
物の存在下でレゾルシンによって生起される比色反応を
用いることにより、スフィンゴ糖脂質中のシアル酸含量
を測定する。Svennerholm's method [Biochem, Biophys, A
cta), Vol. 24 (1957), pp. 604-611], glycosphingolipids are extracted from plasma using a 2:1 mixture of chloroform and methanol, and glycosphingolipids are separated from other lipids in the blood. The sialic acid content in the glycosphingolipids is determined by separating them from the fraction and using a colorimetric reaction generated by resorcinol in the presence of cupric compounds.
カドボディス（に５ｔｏｐｏｄｉｓ） ［米国特許筒４
，３４２゜５６７号明細書］によれば、初め、燐タング
ステン酸を用いて、スフィンゴ糖脂質分画をメタノール
溶液の上清から沈澱させ１次いで、これを再溶解させた
溶液を利用して、比色法による測定を行う。Cadbodis (ni5topodis) [U.S. Patent Tube 4
, 342゜567], first, a glycosphingolipid fraction was precipitated from the supernatant of a methanol solution using phosphotungstic acid, and then a solution obtained by redissolving this was used to Perform colorimetric measurements.
このようにすれば、スパンネルホルム法における特異性
を増大させることができる。しかしながら、シアル酸は
、酸素反応を用いでも測定することができる。In this way, the specificity of the Spannerholm method can be increased. However, sialic acid can also be measured using an oxygen reaction.
クリ二カ・キミ力・アクタ［Ｃ１１ｎ、Ｃｈｉｍ、Ａｃ
ｔａ、 １０８巻（１９８０年）、４９３〜４９８ペー
ジ］に掲載の方法によれば、ノイラミニダーゼなる酵素
を用いて糖蛋白質を分割し、遊離されたシアル酸を、Ｎ
−アセチルノイラミン酸アルドラーゼなる酵素に接触さ
せて、ピルビン酸およびＮ−アセチル−０−マンノサミ
ンを得る。生成したピルビン酸を、ピルビン酸酸化酵素
を用いて酸化させて過酸化物とし、ペルオキシダーゼの
存在下で、Ｐ−クロロフェニル、および４−アミノ−ア
ンチビリンを用いて、これを着色物質に転化する。生成
した着色物質の量を、 ５０５ｎｍの波長にて定量する
。Clinica Kimi Power Acta [C11n, Chim, Ac
TA, Vol. 108 (1980), pp. 493-498], glycoproteins are split using the enzyme neuraminidase, and the liberated sialic acid is converted into N
- Contact with the enzyme acetylneuraminic acid aldolase to obtain pyruvate and N-acetyl-0-mannosamine. The pyruvate produced is oxidized to peroxide using pyruvate oxidase, which is converted to a colored substance using P-chlorophenyl and 4-amino-antivilline in the presence of peroxidase. The amount of colored substance produced is quantified at a wavelength of 505 nm.
上記の方法は、いずれも、測定誤差が２０％を越えると
いう短所が共通している。各著者らによれば、シアル酸
の正常値と異常値との差は非常に小さい（１８ＩＩｇ％
以下は正常と見なされ、 ２０＋ａｇ％を越えるものは
異常と見なされる）ということであるから、その結果は
、悪性疾患の存在を示す可能性はあるが、腫瘍の存在、
あるいはその不在の明白な証明として、これを用いるこ
とはできない、更に、関節炎、乾癖、潰瘍、炎症、＃１
人科疾患など、ある種の良性疾患によっても、血中シア
ル酸含量の増大がもたらされることがある［ジャーナル
・オブ・クリニカル・ケミストリー・アンド・バイオケ
ミストリー（Ｊ 、Ｃｌ１ｎ、Ｃｈｅｍ、Ｂｉｏｃｈｅ
＋ｓ、）、２２巻（１９８４年）。All of the above methods have a common disadvantage of a measurement error of over 20%. According to the authors, the difference between normal and abnormal sialic acid values is very small (18IIg%
20+ag% is considered normal, and anything over 20+ag% is considered abnormal), so the results may indicate the presence of a malignant disease, but they may also indicate the presence of a tumor.
or it cannot be used as clear proof of its absence. Moreover, arthritis, psoriasis, ulcers, inflammation, #1
Certain benign diseases, such as human diseases, can also lead to increased blood sialic acid content [Journal of Clinical Chemistry and Biochemistry (J, Cl1n, Chem, Bioche.
+s, ), vol. 22 (1984).
６４７〜６５１ページコという点も短所である。Another disadvantage is that it is 647 to 651 pages long.
その結果、血中シアル酸含量の測定を目的として現在ま
でに案出された方法は、癌の疑いを生じさせるに過ぎず
、より正確な診断を下すための基盤となり得るものでは
ない。As a result, the methods devised to date for the purpose of measuring blood sialic acid content only raise the suspicion of cancer, and cannot serve as a basis for making a more accurate diagnosis.
本発明の目的は、より信頼性の高い診断をもたらし、か
つ、非常に早期に腫瘍の存在を確認することを可能にし
、また、腫瘍の存在とその不在を互いに区別するための
信頼の置ける情報を提供し、更に、腫瘍の存在する可能
性のある部位の特定に関して、確たる結論を下すことを
可能にするような分析方法を提供することにある。The aim of the invention is to bring about a more reliable diagnosis and make it possible to confirm the presence of a tumor at a very early stage, and also to provide reliable information to distinguish the presence of a tumor from its absence from each other. Furthermore, it is an object of the present invention to provide an analysis method that makes it possible to make a firm conclusion regarding the identification of a site where a tumor may exist.
〔課題を解決するための手段〕
本発明は、 ＰＨ１およびイオン強度が適切であれば、
各種の金属塩の利用によって、血中に存在する糖脂質複
合体の複雑な構成要素を明確に区別される下位分画に分
離することができるとの認識に基づいている。[Means for Solving the Problems] The present invention provides the following: If PH1 and ionic strength are appropriate,
It is based on the recognition that the complex components of glycolipid complexes present in blood can be separated into distinct sub-fractions by the use of various metal salts.
起源の異なる癌腫の進行過程、あるいは、非腫瘍性疾患
において、シアル酸は異なる糖脂質分画に選択的に結合
する。したがって、分離された糖脂質の下位分画におけ
るシアル酸含量を測定し、あるいは、シアル酸が結合し
ている糖脂質の性質を特定することにより、シアル酸含
量の増大が腫瘍の進行によるものか、あるいは（炎症、
関節炎、婦人科疾患などのような）良性の疾病によるも
のかに対する結論を下し、かつ、腫瘍が進行している場
合は、腫瘍の存在する可能性のある部位、および転移の
存在に関して結論を下すことができる。Sialic acid selectively binds to different glycolipid fractions during the progression of cancers of different origins or in non-neoplastic diseases. Therefore, by measuring the sialic acid content in isolated glycolipid subfractions or identifying the nature of the glycolipids to which sialic acid is bound, it is possible to determine whether the increase in sialic acid content is due to tumor progression. , or (inflammation,
Draw a conclusion as to whether the tumor is due to a benign disease (such as arthritis, gynecological disease, etc.) and, if the tumor is advanced, the possible location of the tumor and the presence of metastases. can be lowered.
更に、カドミウム塩を特徴とする特定の金属塩を沈澱剤
として利用した場合、沈澱剤として燐タングステン酸を
用いた場合よりもはるかに早期に、腫瘍状態の認識が可
能であることも観察されている。Furthermore, it has been observed that when certain metal salts, characterized by cadmium salts, are used as precipitants, the tumor status can be recognized much earlier than when phosphotungstic acid is used as the precipitant. There is.
上記の事実に基づき、本発明は、人血を初めとする血液
中の、シアル酸を有する糖脂質複合体の選択的な測定を
目的とする方法に関するものであって、血清を水で希釈
し、次いで、水と混和しない有機溶媒または溶媒混合液
を用いた抽出によって中性脂質を除去する段階と、その
水相から、シアル酸を有する糖脂質が含まれるリボ蛋白
質分画を沈澱させる段階と、沈澱した分画を再溶解させ
る段階と、溶液のシアル酸含有量、あるいは、シアル酸
を有する糖脂質の量および質を、それ自体公知の方法を
用いて測定する段階とで構成される。Based on the above facts, the present invention relates to a method for selectively measuring glycolipid complexes having sialic acid in blood including human blood, which method involves diluting serum with water. , then removing neutral lipids by extraction with a water-immiscible organic solvent or solvent mixture, and precipitating a riboprotein fraction containing sialic acid-bearing glycolipids from the aqueous phase. , a step of redissolving the precipitated fraction, and a step of measuring the sialic acid content of the solution, or the quantity and quality of sialic acid-containing glycolipids, using methods known per se.
本発明によれば、沈澱剤として、混合液中の最終金属イ
オン濃度がｏ、ｏｏｓ〜０．１モル／２に対応する量の
、アルカリ金属以外の１価、２価、あるいは３価の水溶
性金属塩を用い、かつ、ｐＨ１．５〜８．０にて沈澱を
生成させる。According to the present invention, as a precipitant, a monovalent, divalent, or trivalent aqueous solution other than an alkali metal is used in an amount corresponding to a final metal ion concentration of o, oos to 0.1 mol/2 in the mixed solution. A precipitate is formed using a neutral metal salt and at a pH of 1.5 to 8.0.
水で希釈した血清から中性脂質を除去するには。To remove neutral lipids from serum diluted with water.
米国特許第４，３４２，５６７号明細書に記載の方法を
適用することが好適である。これによれば、抽出剤とし
て、クロロホルムとメタノールの体積比２：１の混合液
を使用する。メタノール水溶液となった上清（以下、メ
タノール溶液の上清と称する）に対しては、更に沈澱過
程を進め、一方、中性脂質のクロロホルム溶液である下
方の相は棄却する。It is preferred to apply the method described in US Pat. No. 4,342,567. According to this, a mixed solution of chloroform and methanol in a volume ratio of 2:1 is used as an extractant. The supernatant, which has become an aqueous methanol solution (hereinafter referred to as the supernatant of the methanol solution), is further subjected to the precipitation process, while the lower phase, which is a chloroform solution of neutral lipids, is discarded.
その後、メタノール水溶液の相、あるいはそのアリコー
トに、ｏ、ｏｏｓ〜０．１モル／Ｑの金属イオンを含む
混合液が得られるような量の沈澱剤を添加する。沈澱剤
は、水溶液として用いるのが好適である。金属イオンの
いくつかに関して、沈澱の際の好適なｐＨ値を以下の表
に示す。Thereafter, a precipitant is added to the aqueous methanol phase, or an aliquot thereof, in an amount such that a mixture containing o, oos to 0.1 mol/Q of metal ions is obtained. The precipitant is preferably used as an aqueous solution. For some metal ions, suitable pH values during precipitation are shown in the table below.
（以下余白）
倉１０膨（Ｚ 匹
第−鉄イオン（Ｆｅ” ”） ３．０〜３．５
第二鉄イオン（Ｆｅ”） ３．０〜５．０コバ
ルトイオン（Ｃｏ″＋）４．０〜５．５ニツケルイオン
（Ｎｉ”） ４．５〜５．５カドミウムイオン（
Ｃｄ”） ４．５〜６＃０亜鉛イオン（Ｚｎ”）
１．５〜５．０銀イオン（Ａｇつ
６．θ〜８．０第８．銀イオン（Ｈｇ”）
１．５〜３．０第二銅イオン（Ｃｕ″＋） ３
．０〜４．５カルシウムイオン（Ｃａ”） ４．５
〜５．５マグネシウムイオン（Ｍｇ”） ５．５〜７
．０その結果生じる沈澱を分離し９次いで。(Left below) Kura 10 volumes (Z ferrous ions (Fe"") 3.0 to 3.5
Ferric ion (Fe") 3.0-5.0 Cobalt ion (Co"+) 4.0-5.5 Nickel ion (Ni") 4.5-5.5 Cadmium ion (
Cd") 4.5~6#0 Zinc ion (Zn")
1.5 to 5.0 silver ions (Ag
6. θ~8.0 8th. Silver ion (Hg”)
1.5-3.0 cupric ion (Cu″+) 3
．． 0-4.5 Calcium ion (Ca”) 4.5
~5.5 Magnesium ion (Mg”) 5.5~7
．． The resulting precipitate was then separated.
（イ）米国特許第４，３４２，５６７号明細書に記載の
、無機酸、第二銅塩、およびレゾルシンを用いて行われ
る比色反応に基づくような公知の方法、あるいは、クリ
二カ・キミ力・アクタ、１０８巻（１９８０年）、 ４
９３〜４９８ページに記載された酵素反応による方法を
用いて、沈澱物中のシアル酸含量を測定するか、あるい
は、
（ロ）クリニカル・ケミストリー（Ｃ１ｉｎ、Ｃｈｅ曽
、）、２２巻（１９７６年）、１，５１６〜１，５２１
ページに記載の高圧液体クロマトグラフィーによるなど
の公知の方法を用いて、シアル酸に結合した糖脂質の種
類および量を測定する。(a) Known methods such as those described in U.S. Pat. No. 4,342,567 and based on a colorimetric reaction performed using an inorganic acid, a cupric salt, and resorcinol; Kimiriki Acta, Volume 108 (1980), 4
Measure the sialic acid content in the precipitate using the enzymatic reaction method described on pages 93 to 498, or (b) Clinical Chemistry (C1in, Che Zeng), Vol. 22 (1976). , 1,516-1,521
The type and amount of glycolipids bound to sialic acid are determined using known methods, such as by high pressure liquid chromatography as described on page 12.
起源を同じくする同一の血液試料のシアル酸含量を、少
なくとも５種類の異なる金属イオンを用いて逐次的に測
定し、正常値に対する測定値の比（「正常値」なる用語
は、所定の金属イオンを沈澱剤として用いて統計的に計
算された、病的でないシアル酸濃度のことである。）の
総和を求めた場合１本発明による方法の鑑別診断値は著
しく増大する。The sialic acid content of the same blood sample of the same origin is measured sequentially using at least five different metal ions, and the ratio of the measured value to the normal value (the term "normal value" refers to the ratio of the measured value to the normal value) The differential diagnostic value of the method according to the invention increases significantly when the sum of the statistically calculated non-pathological sialic acid concentrations using 1 as a precipitant is determined.
発明者らの経験によれば、Σ（測定値／正常値）の値は
、（局在する腫瘍に関しては）次の順に減少する。すな
わち、乳腫瘍−結腸腫瘍−卵巣腫瘍一頭部または頚部腫
瘍−脇癌一非腫瘍性疾病一腫瘍のない健康状態の順であ
る。According to our experience, the value of Σ(measured value/normal value) decreases (for localized tumors) in the following order: That is, the order is breast tumor - colon tumor - ovarian tumor - head or neck tumor - armpit cancer - non-neoplastic disease - tumor-free health status.
同様に１本発明に従って、少なくとも５種類の異なる金
属イオンを用いて行なったシアル酸の測定結果を、公知
のシアル酸の測定結果（例えば。Similarly, according to the present invention, the results of measurements of sialic acid carried out using at least five different metal ions can be compared with the results of measurements of known sialic acids (e.g.
その起源に関係なくシアル酸の含量を測定した。The content of sialic acid was determined regardless of its origin.
米国特許第４，３４２，５６７号明細書に記載の測定結
果など）で除し、これらの商の総和を求めると、鑑別診
断級数が得られる０個々の商からも、鑑別診断値を求め
ることができる。When dividing by the measurement results described in U.S. Pat. No. 4,342,567, etc.) and calculating the sum of these quotients, a differential diagnostic series is obtained.Differential diagnostic values can also be calculated from the individual quotients. Can be done.
以下１本発明を制約するものではない実施例を援用して
１本発明の詳細な説明する。Hereinafter, the present invention will be described in detail with reference to examples that do not limit the present invention.
実施例において行われた。血液試料に対する前処理、お
よび、沈澱物中のシアル酸含量の測定方法は下記のとお
りである。This was done in the example. The pretreatment for the blood sample and the method for measuring the sialic acid content in the precipitate are as follows.
測定には静脈血を用いる。採取後の血液試料に。Venous blood is used for measurement. For blood samples after collection.
４．０００ｒ、ｐ、ｓ、の遠心分離を１５分間行い、試
料０．６＋ｗｌを凍結させる。試料中の脂質に結合した
シアル酸の濃度は、−２０℃で貯蔵すれば６ケ月間変化
することがない。Centrifuge at 4.000 r, p, s for 15 minutes and freeze the sample 0.6+wl. The concentration of lipid-bound sialic acid in a sample remains unchanged for 6 months when stored at -20°C.
分析開始に先立ち、メタノールを用いて試験管を完全に
洗浄し、乾燥し、冷凍庫に入れて冷却しておく、この試
験管それぞれに、血清試料０．２ａ＋１、および二回蒸
留水０．２ｍｌを注入する。Before starting the analysis, the test tubes were thoroughly washed with methanol, dried, and kept cool in the freezer.To each test tube was added 0.2a+1 serum sample and 0.2ml of double-distilled water. inject.
混合液を、ポルテックス（Ｖｏｒｔｅｘ）攪拌器を用い
て１５分間攪拌し、次いで、氷冷水浴（０℃）中に放置
する。被験血清および対照群のそれぞれから、平行試料
として各２例を用いる。The mixture is stirred for 15 minutes using a Vortex stirrer and then placed in an ice-cold water bath (0° C.). Two parallel samples are used from each of the test serum and control groups.
予め冷却しておいたクロロホルムとメタノールの体積比
２：１の混合液３ｍｌを、すべての試験管に注入し、試
験管の内容を、ポルテックス攪拌器を用いて３０秒間攪
拌する６次いで、再び試験管を氷冷水浴中に静置する。Inject 3 ml of a pre-cooled mixture of chloroform and methanol in a volume ratio of 2:1 into all test tubes and stir the contents of the test tubes for 30 seconds using a Portex stirrer6.Then test again. Place the tube in an ice-cold water bath.
各試料に、予め＋４℃に冷却しておいた二回蒸留水０．
５閣１を加え、試験管の内容を、ポルテックス攪拌器を
用いて１５秒間攪拌する０次いで、混合液を室温にて５
分間静置する。Add 0.0 ml of double-distilled water to each sample, pre-cooled to +4°C.
Add 5 parts and stir the contents of the test tube for 15 seconds using a Portex stirrer.Then, stir the mixture at room temperature for 5 seconds.
Let stand for a minute.
その後、試料に３．００Ｏｒ、ｐ、■、の遠心分離を１
０分間行う、各上清１ｍｌを１番号を付した遠沈管に注
意深く注入し、各実施例中に示した量および濃度の金属
塩水溶液を加え、混合液のＰＨを各実施例中に示した値
に調整し、各遠沈管の内容を、ポルテックス攪拌器を用
いて攪拌し、次いで、５分間静置する。After that, the sample was centrifuged at 3.00 Or, p, 1
Carefully pour 1 ml of each supernatant into centrifuge tubes numbered 1, add metal salt aqueous solution in the amount and concentration indicated in each example, and add the PH of the mixture as indicated in each example. The contents of each centrifuge tube are stirred using a Portex stirrer and then allowed to stand for 5 minutes.
静置期間の終了時に、各遠沈管の内容に４．０００ｒ、
ｐ、ｍ、の遠心分離を１５分間行い１次いで、上清を静
かに流出させて沈澱から完全に分離する（上清は棄却す
る）。At the end of the standing period, add 4,000 r to the contents of each centrifuge tube.
Perform centrifugation at P and M for 15 minutes, then gently drain the supernatant to separate it completely from the precipitate (discard the supernatant).
トリス緩衝液１■ｌを各遠沈管に注入して、沈澱をこれ
に再溶解させる。トリス緩衝液は、トリス（ヒドロキシ
メチル）アミノメタン１０ミリモルを。Inject 1 liter of Tris buffer into each centrifuge tube to redissolve the precipitate therein. Tris buffer contains 10 mmol of tris(hydroxymethyl)aminomethane.
二回蒸留水１１２に溶解させ、溶液のｐＨを８．６に整
えて調製したものである。It was prepared by dissolving it in double distilled water 112 and adjusting the pH of the solution to 8.6.
光度測定用の標準試料および盲検試料の調製法は、下記
のとおりである。The method for preparing standard samples and blind samples for photometric measurements is as follows.
分析化学的に純粋なシアル酸５０■を、二回蒸留水１０
０ｍ１に溶解させ、この溶液をストック液として、二回
蒸留水で更に２回希釈する（濃度は２５■７１００醜１
、および１２．５■／１００■ｌとなる）。Analytically, 50 μl of chemically pure sialic acid was added to 10 μl of double-distilled water.
Use this solution as a stock solution and dilute it twice with double distilled water (concentration is 25 x 7100 ugly 1).
, and 12.5■/100■l).
標準溶液を、＋４℃にて貯蔵する。この３種類の濃度の
標準溶液０．２ｍｌを試料として遠沈管に注入し、二回
蒸留水を用いて、各遠沈管内の溶液の体積を１■ｌに調
整する。盲検試料としては、二回蒸留水１■１を用いる
。Standard solutions are stored at +4°C. Inject 0.2 ml of the standard solutions of these three concentrations into centrifuge tubes as samples, and adjust the volume of the solution in each centrifuge tube to 1 liter using double-distilled water. As a blind sample, 1×1 portion of double distilled water is used.
レゾルシンなる試薬の溶液各１■１を、すべての試料（
ＩＩ準試料、盲検試料、対照試料、被験血清）に添加す
る。試薬用レゾルシン溶液の調製法は。Add 1 x 1 solution of resorcin reagent to all samples (
II quasi-sample, blind sample, control sample, test serum). How to prepare resorcinol solution for reagents.
下記のとおりである。It is as follows.
分析化学的に純粋なレゾルシン２ｇを、二回蒸留水１０
０ｍ１に溶解させる。無水硫酸第二銅２．４９ｇを。2 g of analytically pure resorcinol in 10 g of double distilled water
Dissolve in 0ml. 2.49 g of anhydrous cupric sulfate.
二回蒸留水１００ｍ１に溶解させる。その後、レゾルシ
ン溶液１０ｍ１を、硫酸第二銅溶液０．２５社、および
二回蒸留水９．７５ｍ１と混合し１分析化学的に純粋な
濃塩酸水溶液１００■ｌを、この混合液に加える。Dissolve in 100 ml of double-distilled water. Thereafter, 10 ml of resorcinol solution are mixed with 0.25 ml of cupric sulfate solution and 9.75 ml of double-distilled water, and 100 ml of analytically pure concentrated aqueous hydrochloric acid solution are added to this mixture.
試験管を、１００℃の水浴中に正確に１０分間放置する
。その後、すべての試験管を、直ちに０℃の氷冷水浴中
に浸し二そのまま１０分間放置する。ブチル酢酸とｎ−
ブタノールの体積比８５：１５の混合液を、２ｍｌずつ
各試験管に注入し、試験管の内容を、ポルテックス攪拌
器を用いて５分間攪拌する。混合液を時折り攪拌しつつ
放置し、試料に２，５００ｔ’、Ｐ、■。Place the test tube in a 100°C water bath for exactly 10 minutes. Thereafter, all test tubes are immediately immersed in an ice-cold water bath at 0° C. and left for 10 minutes. Butylacetic acid and n-
2 ml of a 85:15 by volume mixture of butanol is injected into each test tube and the contents of the test tubes are stirred for 5 minutes using a Portex stirrer. The mixed solution was left to stand with occasional stirring, and 2,500 t', P, ■ was applied to the sample.
の遠心分離を１０分間行い、次いで、青色を呈するその
上清を、各遠沈管から測定用キュベツトに移す、この青
色の濃さは、６か月以内に変化することはない。centrifugation for 10 minutes and then transfer the supernatant, which has a blue color, from each centrifuge tube to a measuring cuvette; the intensity of this blue color does not change within 6 months.
光度測定は、５８０ｎ＋ｍの波長を用いて行う、二回蒸
留水に合わせて、光度計のゼロ点を調整する。Photometric measurements are performed using a wavelength of 580n+m, zeroing the photometer for double-distilled water.
その後、光度計の（単一光線または二重光線を用いる）
型式に応じて、盲検試料に対する測定を行い、あるいは
、下式により、盲検試料の吸光度を、測定された被験試
料の吸光度から減じるなどして。Then the photometer (using single beam or double beam)
Depending on the model, measurements are performed on a blind sample, or the absorbance of the blind sample is subtracted from the measured absorbance of the test sample using the formula below.
補正値を求める。Find the correction value.
補正された吸光度＝測定された吸光度−盲検試料の吸光
度
シアル酸含量は、下記の２種類の方法のいずれかを用い
て算出する。Corrected absorbance=measured absorbance−absorbance of blind sample Sialic acid content is calculated using either of the following two methods.
（１）既知のシアル酸濃度の標準シアル酸試料を用いて
得られた結果から、吸光度対濃度の曲線が。(1) An absorbance versus concentration curve from the results obtained using a standard sialic acid sample of known sialic acid concentration.
グラフ上、あるいは計算によって求められる。Determined on a graph or by calculation.
希釈の段階が存在するため、以下の表に従い。As there are stages of dilution, follow the table below.
本来の濃度を再計算する必要がある。It is necessary to recalculate the original concentration.
（以下余白）
本来の濃度 測定された濃度 吸光度
（＋ｓｇ／１００ｍ１） （ｆｉ ｇ／２００
ｍ１）ｓｏ ｉｏｏ ｏ、
ｓｏｇ２５ ５０ ０．２５２１
２．５ ２５ ０．１２７上記に
従って作成した較正曲線から１体積１ｍｌ中の上清試料
のシアル酸含量（Ａ）を決定することができる。この曲
線は、上清１ｍｌあたりの＃ｇで表したシアル酸の量を
単位とするシアル酸含量を示す、血清中のシアル酸含量
は１式シアル酸含量（ｍｇ／１００ｍ１）＝ＡＸ Ｏ，
７２（式中、Ａは、 Ｉｔｇ／Ｉｌｌを単位とする上溝
中のシアル酸含量を意味し、０．７２は、希釈度と、ｊ
ｌｇがら■への換算率とから成る係数である。）から算
出することができる。(Left below) Original concentration Measured concentration Absorbance (+sg/100m1) (fig/200
m1) so ioo o,
sog25 50 0.2521
2.5 25 0.127 The sialic acid content (A) of the supernatant sample in a volume of 1 ml can be determined from the calibration curve prepared according to the above. This curve shows the sialic acid content in units of the amount of sialic acid expressed in #g per ml of supernatant.
72 (where A means the sialic acid content in the upper groove in Itg/Ill, 0.72 is the dilution degree, and j
This is a coefficient consisting of the conversion rate from lg to ■. ) can be calculated from
（２）第２の計算方法においては、既知のシアル酸濃度
の参照用血清試料を用いる。数回の平行測定から、式
％式％（）
（式中、Ｃ（ｒｅｆ）は、＠／１００ｍ１で表された参
照用血清のシアル酸含量を、Ｅ（ｒｅｆ）は、参照用血
清の吸光度をそれぞれ意味する。）
によって係数Ｆを算出する。被験試料のシアル酸含量は
、式
％式％
（式中、Ｅ（ｍ）は、被験試料において測定された吸光
度を意味する。）
から算出することができる。(2) In the second calculation method, a reference serum sample with a known sialic acid concentration is used. From several parallel measurements, the formula % formula % () (where C(ref) is the sialic acid content of the reference serum expressed in @/100ml, and E(ref) is the absorbance of the reference serum respectively.) Calculate the coefficient F by The sialic acid content of the test sample can be calculated from the formula % (where E(m) means the absorbance measured in the test sample).
失産叢上
健康な人間から採取した血清に、前述のクロロホルムと
メタノールの混合液を用いた抽出を行う。Serum collected from a healthy human being undergoing abortifacients is extracted using the above-mentioned mixture of chloroform and methanol.
ｐＨ５，５で、メタノール溶液の上清と、塩化ニッケル
の０．５モル／Ω溶液とを１０：１の比率で混合して沈
澱を生成させ、沈澱中のシアル酸含量を測定する。 ４
．６５■Ｘなる値が得られた。At pH 5.5, the supernatant of the methanol solution and a 0.5 mol/Ω solution of nickel chloride are mixed at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured. 4
．． A value of 65■X was obtained.
ス〕１剋」工
健康な人間から採取した血清に、前記クロロホルムとメ
タノール、の混合液を用いた抽出を行う。1) Extraction is performed on serum collected from a healthy human using the mixture of chloroform and methanol.
ｐＨ５．３で、メタノール溶液の上清と、塩化ニッケル
の１．０モル／Ω溶液とを１０＝１の比率で混合して沈
澱を生成させ、沈澱中のシアル酸含量を測定する。 ６
．４３■ヌなる値が得られた。At pH 5.3, the supernatant of the methanol solution and a 1.0 mol/Ω solution of nickel chloride are mixed at a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured. 6
．． A value of 43■nu was obtained.
失凰涯ｌ
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Loss of Death: Serum collected from a healthy human is extracted using the above-mentioned mixture of chloroform and methanol.
ＰＨ５，９で、メタノール溶液の上清と、塩化カドミウ
ムの０．１モル／Ω溶液とを１０：１の比率で混合して
沈澱を生成させ、沈澱中のシアル酸含量を測定する。
ｉｔ、ｓｚ■算なる値が得られた。At pH 5.9, the supernatant of the methanol solution and a 0.1 mol/Ω solution of cadmium chloride are mixed at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
The following values were obtained: it, sz.
寒凰叢土
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Kano Sodo: Serum collected from a healthy human is extracted using the mixture of chloroform and methanol.
ｐＨ５，５で、メタノール溶液の上清と、塩化カドミウ
ムの０．５モル／Ω溶液とをｌＯ：１の比率で混合して
沈澱を生成させ、沈澱中のシアル酸含量を測定する。
１０．４８■％なる値が得られた。At pH 5.5, the supernatant of the methanol solution and a 0.5 mol/Ω solution of cadmium chloride are mixed at a ratio of 1O:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
A value of 10.48% was obtained.
ヌ」１匹」−
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。``1 animal'' - Extraction is performed on serum collected from a healthy human using the above-mentioned mixture of chloroform and methanol.
ｐ）１５．３で、メタノール溶液の上滑と、塩化カドミ
ウムの１．０モル／Ω溶液とを１０：１の比率で混合し
て沈澱を生成させ、沈澱中のシアル酸含量を測定する。p) At 15.3, a precipitate is formed by mixing the methanol solution and a 1.0 mol/Ω solution of cadmium chloride in a ratio of 10:1, and the sialic acid content in the precipitate is measured.
１４．８１■％なる値が得られた。A value of 14.81% was obtained.
寒五匠且
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from Kangosho and a healthy person is extracted using the above-mentioned mixture of chloroform and methanol.
ｐＨ３，３で、メタノール溶液の上滑と、硫酸第一鉄の
０．１モル／Ω溶液とをｌ０＝１の比率で混合して沈澱
を生成させ、沈澱中のシアル酸含量を測定する。At pH 3.3, a precipitate is formed by mixing a methanol solution and a 0.1 mol/Ω solution of ferrous sulfate at a ratio of l0=1, and the sialic acid content in the precipitate is measured.
９．０８．％なる値が得られた。9.08. A value of % was obtained.
寒産叢ユ
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a healthy human being is extracted using the mixture of chloroform and methanol.
ｐＨ３．０で、メタノール溶液の上清と、硫酸第一鉄の
０．５モル／Ω溶液とをｌＯ：１の比率で混合して沈澱
を生成させ、沈澱中のシアル酸含量を測定する。At pH 3.0, the supernatant of the methanol solution and a 0.5 mol/Ω solution of ferrous sulfate are mixed at a ratio of 1O:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
９．５４．％なる値が得られた。9.54. A value of % was obtained.
ヌＪＬ糎」−
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。``Nu JL Glue'' - Serum collected from a healthy human is subjected to extraction using the above-mentioned mixture of chloroform and methanol.
ｐＨ２，９で、メタノール溶液の上清と、硫酸第一鉄の
１．０モル／Ω溶液とを１０＝１の比率で混合して沈澱
を生成させ、沈澱中のシアル酸含量を測定する。At pH 2.9, the supernatant of the methanol solution and a 1.0 mol/Ω solution of ferrous sulfate are mixed at a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured.
９．５２．％なる値が得られた。9.52. A value of % was obtained.
失五五主
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a healthy human being is extracted using the mixture of chloroform and methanol.
ｐＨ２，７で、メタノール溶液の上清と、塩化亜鉛の０
．１モル／Ω溶液とを１０：１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。At pH 2.7, supernatant of methanol solution and zinc chloride
．． 1 mol/Ω solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
６．５２ｍｇ％なる値が得られた。A value of 6.52 mg% was obtained.
失凰舊烈
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a healthy human being is extracted using the mixture of chloroform and methanol.
ｐＨ１．９で、メタノール溶液の上清と、塩化亜鉛の０
．５モル／Ω溶液とをｌＯ：１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。At pH 1.9, the methanol solution supernatant and zinc chloride
．． 5 mol/Ω solution at a ratio of lO:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
１２．３４■％なる値が得られた。A value of 12.34% was obtained.
末凰叢旦
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a healthy human being is extracted using the mixture of chloroform and methanol.
ＰＨ４，４で、メタノール溶液の上清と、硝酸鉛の０．
１モル／１２溶液とをｌＯ：１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。At pH 4.4, the supernatant of methanol solution and 0.00% of lead nitrate.
A 1 mol/12 solution is mixed at a ratio of 1O:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
８．６１ｍｇ％なる値が得られた。A value of 8.61 mg% was obtained.
叉直涯Ｕ
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Extraction is performed on serum collected from a healthy human using the mixture of chloroform and methanol.
ｐ）１３．７で、メタノール溶液の上清と、硝酸鉛の０
．５モル／Ｑ溶液とを１０＝１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。p) At 13.7, the supernatant of the methanol solution and the 0% of lead nitrate
．． 5 mol/Q solution is mixed at a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured.
１２．１１１１ｇ％なる値が得られた。A value of 12.1111 g% was obtained.
叉産五Ｎ
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。5N Serum collected from a healthy human is extracted using the above-mentioned mixture of chloroform and methanol.
ｐＨ３．５で、メタノール溶液の上清と、硝酸鉛の１．
０モルＩＱ溶液とをｌＯ：１の比率で混合して沈澱を生
成させ、沈澱中のシアル酸含量を測定する。 １２．３
５ＩＩＩｇ％なる値が得られた。At pH 3.5, the supernatant of the methanol solution and 1.
A precipitate is formed by mixing with a 0 molar IQ solution at a ratio of 1O:1, and the sialic acid content in the precipitate is measured. 12.3
A value of 5IIIg% was obtained.
叉五透旦
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a healthy human is extracted using the mixture of chloroform and methanol.
ＰＨ８，５で、メタノール溶液の上清と、硝酸銀の０．
１モル／Ｑ溶液とを１０＝１の比率で混合して沈澱を生
成させ、沈澱中のシアル酸含量を測定する。 ９．８９
■％なる値が得られた。At pH 8.5, the supernatant of methanol solution and 0.0.
1 mol/Q solution is mixed at a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured. 9.89
■A value of % was obtained.
失産涯耗
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a healthy human being is extracted using the above-mentioned mixture of chloroform and methanol.
ｐＨ６，０で、メタノール溶液の上清と、硝酸銀の０．
５モル／Ｑ溶液とを１０＝１の比率で混合して沈澱を生
成させ、沈澱中のシアル酸含量を測定する。 １４．６
７■％なる値が得られた。At pH 6.0, supernatant of methanol solution and 0.0.
5 mol/Q solution is mixed at a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured. 14.6
A value of 7% was obtained.
寒遼■圧
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Blood serum collected from a healthy human is extracted using the mixture of chloroform and methanol.
ｐＨ５，８で、メタノール溶液の上清と、硝酸銀の１．
０モル／Ｑ溶液とを１０：１の比率で混合して沈澱を生
成させ、沈澱中のシアル酸含量を測定する。 １４．８
１■％なる値が得られた。At pH 5.8, supernatant of methanol solution and 1.
0 mol/Q solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured. 14.8
A value of 1% was obtained.
スｍ
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Sm Extraction is performed on serum collected from a healthy human using the mixture of chloroform and methanol.
ｐＨ１．９で、メタノール溶液の上清と、塩化第二水銀
の０．１モル／ｊ２溶液とを１０：１の″比率で混合し
て沈澱を生成させ、沈澱中のシアル酸含量を測定する。At pH 1.9, the supernatant of the methanol solution and the 0.1 mol/j2 solution of mercuric chloride are mixed in a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured. .
６．０５■％なる値が得られた。A value of 6.05% was obtained.
失凰叢圏
健康な人間から採取した卑情に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Extraction using the above-mentioned mixture of chloroform and methanol is performed on the phlegm collected from a healthy human.
ｐＨ１．７で、メタノール溶液の上清と、塩化第二水銀
の０．５モル／ρ溶液とを１０＝１の比率で混合して沈
澱を生成させ、沈澱中のシアル酸含量を測定する。 ７
．６８ｍｇ％なる値が得られた。At pH 1.7, the supernatant of the methanol solution and the 0.5 mol/ρ solution of mercuric chloride are mixed in a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured. 7
．． A value of 68 mg% was obtained.
失産五皿
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a healthy human being with a miscarriage is subjected to extraction using the above-mentioned mixture of chloroform and methanol.
ＰＩ（１，５で、メタノフル溶液の上清と、塩化第二水
銀の１．０モルＩＱ溶液とを１０＝１の比率で混合して
沈澱を生成させ、沈澱中のシアル酸含量を測定する。
１１．９９■％なる値が得られた。PI (at 1,5), mix the supernatant of the methanoflu solution and a 1.0 molar IQ solution of mercuric chloride in a ratio of 10=1 to form a precipitate, and measure the sialic acid content in the precipitate. .
A value of 11.99% was obtained.
失直涯毀
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a delinquent and healthy human is subjected to extraction using the mixture of chloroform and methanol.
ＰＨ４，２で、メタノール溶液の上清と、硫酸第二銅の
０．１モルＩＱ溶液とをｌＯ：１の比率で混合して沈澱
を生成させ、沈澱中のシアル酸含量を測定する。At pH 4.2, the supernatant of the methanol solution and a 0.1 molar IQ solution of cupric sulfate are mixed in a ratio of 1O:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
１０．１３■％なる値が得られた。A value of 10.13% was obtained.
尖凰五旦
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a healthy human is extracted using the mixture of chloroform and methanol.
ｐＨ３．８で、メタノール溶液の上清と、硫酸第二銅の
０．５モルＩＱ溶液とを１０：１の比率で混合して沈澱
を生成させ、沈澱中のシアル酸含量を測定する。At pH 3.8, the supernatant of the methanol solution and a 0.5 molar IQ solution of cupric sulfate are mixed in a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
１０．４８■％なる値が得られた。A value of 10.48% was obtained.
失五叢Ｕ
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Loss of Pentaplexus U Serum collected from a healthy human is extracted using the above-mentioned mixture of chloroform and methanol.
ｐ）１３．６で、メタノール溶液の上清と、硫酸第二銅
の１．０モルＩＱ溶液とをｌＯ：１の比率で混合して沈
澱を生成させ、沈澱中のシアル酸含量を測定する。p) At 13.6, mix the supernatant of the methanol solution and a 1.0 molar IQ solution of cupric sulfate in a ratio of lO:1 to form a precipitate, and measure the sialic acid content in the precipitate. .
１２．２２Ｒ％なる値が得られた。A value of 12.22R% was obtained.
失産五翌
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a healthy human being five days after miscarriage is extracted using the mixture of chloroform and methanol.
ｐ）１５．５で、メタノール溶液の上清と、塩化カルシ
ウムの０．１モル／Ｑ溶液とを１０＝１の比率で混合し
て沈澱を生成させ、沈澱中のシアル酸含量を測定する。p) At 15.5, the supernatant of the methanol solution and the 0.1 mol/Q solution of calcium chloride are mixed in a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured.
３．７２■％なる値が得られた。A value of 3.72% was obtained.
失１匠旦
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a healthy human is extracted using the mixture of chloroform and methanol.
ｐＨ５，４で、メタノール溶液の上溝と、塩化カルシウ
ムの０．５モルＩＱ溶液とをｌＯ：１の比率で混合して
沈澱を生成させ、沈澱中のシアル酸含量を測定する。３
．０２■％なる値が得られた。At pH 5.4, a precipitate is formed by mixing a methanol solution and a 0.5 molar IQ solution of calcium chloride in a ratio of 1O:1, and the sialic acid content in the precipitate is measured. 3
．． A value of 0.02% was obtained.
失五■並
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a normal and healthy human being is extracted using the above-mentioned mixture of chloroform and methanol.
ｐ）１５．３で、メタノール溶液の上清と、塩化カルシ
ウムの１．０モルＩＱ溶液とを１０：１の比率で混合し
て沈澱を生成させ、沈澱中のシアル酸含量を測定する。p) At 15.3, mix the supernatant of the methanol solution and a 1.0 molar IQ solution of calcium chloride in a ratio of 10:1 to form a precipitate and measure the sialic acid content in the precipitate.
７．０５■％なる値が得られた。A value of 7.05% was obtained.
寒１涯垣
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a healthy human being is extracted using the above-mentioned mixture of chloroform and methanol.
ｐＨ６，１で、メタノール溶液の上清と、塩化マグネシ
ウムの０．１モル／２溶液とを１０：１の比率で混合し
て沈澱を生成させ、沈澱中のシアル酸含量を測定する。At pH 6.1, the supernatant of the methanol solution and a 0.1 mol/2 solution of magnesium chloride are mixed at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
４．１９＋ａｇ％なる値が得られた。A value of 4.19+ag% was obtained.
叉凰盤互
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a healthy human is extracted using the mixture of chloroform and methanol.
ＰＨ６，７で、メタノール溶液の上清と、塩化マグネシ
ウムの０．５モル／Ｑ溶液とを１０；１の比率で混合し
て沈澱を生成させ、沈澱中のシアル酸含量を測定する。At pH 6.7, the supernatant of the methanol solution and a 0.5 mol/Q solution of magnesium chloride are mixed at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
３．６１■算なる値が得られた。A value of 3.61 mm was obtained.
失産叢旦
健康な人間から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a healthy human being who has suffered a miscarriage is extracted using the above-mentioned mixture of chloroform and methanol.
ｐ）１６．５で、メタノール溶液の上清と、塩化マグネ
シウムの１．０モル／Ｑ溶液とを１０＝１の比率で混合
して沈澱を生成させ、沈澱中のシアル酸含量を測定する
。 ４．８１１１Ｉｇ％なる値が得られた。p) At 16.5, the supernatant of the methanol solution and the 1.0 mol/Q solution of magnesium chloride are mixed in a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured. A value of 4.8111 Ig% was obtained.
失五五並
乳癌患者から採取した血清に、前記クロロホルムとメタ
ノールの混合液を用いた抽出を行う。Serum collected from a breast cancer patient is extracted using the mixture of chloroform and methanol.
ｐＨ５，３で、メタノール溶液の上清と、塩化ニッケル
の１．０モル／Ｑ溶液とをｌＯ：１の比率で混合して沈
澱を生成させ、沈澱中のシアル酸含量を測定する。 ２
８．８７ｍｇ％なる値が得られた。At pH 5.3, the supernatant of the methanol solution and a 1.0 mol/Q solution of nickel chloride are mixed at a ratio of 1O:1 to form a precipitate, and the sialic acid content in the precipitate is measured. 2
A value of 8.87 mg% was obtained.
失１舊赳
乳癌患者から採取した血清に、前記クロロホルムとメタ
ノールの混合液を用いた抽出を行う。Serum collected from a patient with breast cancer is extracted using the mixture of chloroform and methanol.
ＰＨ５，３で、メタノール溶液の上清と、塩化カドミウ
ムの１．０モル／Ｑ溶液とを１０：１の比率で混合して
沈澱を生成させ、沈澱中のシアル酸含量を測定する。
４１，３１■Ｘなる値が得られた。At pH 5.3, the supernatant of the methanol solution and a 1.0 mol/Q solution of cadmium chloride are mixed at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
A value of 41,31*X was obtained.
失凰亘旦
乳癌患者から採取した血清に、前記クロロホルムとメタ
ノールの混合液を用いた抽出を行う。Serum collected from a patient with breast cancer is extracted using the mixture of chloroform and methanol.
ｐＨ２，９で、メタノール溶液の上清と、硫酸第一鉄の
１．０モル／ｊ１溶液とを１０＝１の比率で混合して沈
澱を生成させ、沈澱中のシアル酸含量を測定する。At pH 2.9, the supernatant of the methanol solution and a 1.0 mol/j1 solution of ferrous sulfate are mixed at a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured.
３３．３９■％なる値が得られた。A value of 33.39% was obtained.
失五涯用
乳癌患者から採取した血清に、前記クロロホルムとメタ
ノールの混合液を用いた抽出を行う。Serum collected from a terminal breast cancer patient is extracted using the above-mentioned mixture of chloroform and methanol.
ｐＨ１，７で、メタノール溶液の上清と、塩化亜鉛の１
．０モル／Ｑ溶液とを１０：１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。At pH 1.7, the supernatant of methanol solution and 1 of zinc chloride
．． 0 mol/Q solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
３９．４１Ｎ％なる値が得られた。A value of 39.41 N% was obtained.
失蓬豊川
乳癌患者から採取した血清に、前記クロロホルムとメタ
ノールの混合液を用いた抽出を行う。Serum collected from a Toyokawa breast cancer patient is extracted using the mixture of chloroform and methanol.
ｐＨ１，７で、メタノール溶液の上清と、硫酸亜鉛の１
．０モル／Ｑ溶液とを１０：１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。At pH 1.7, the supernatant of methanol solution and 1 of zinc sulfate
．． 0 mol/Q solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
３ｇ、９０．％なる値が得られた。3g, 90. A value of % was obtained.
失五五旦
乳癌患者から採取した血清に、前記クロロホルムとメタ
ノールの混合液を用いた抽出を行う。Serum collected from a patient with breast cancer is extracted using the mixture of chloroform and methanol.
ｐＨ３，５で、メタノール溶液の上滑と、硝酸鉛の１．
０モルＩＱ溶液とを１０：１の比率で混合して沈澱を生
成させ、沈澱中のシアル酸含量を測定する。 ４５．０
２ｍｇ％なる値が得られた。At pH 3.5, add a methanol solution and 1.0 g of lead nitrate.
A precipitate is formed by mixing with a 0 molar IQ solution in a ratio of 10:1, and the sialic acid content in the precipitate is measured. 45.0
A value of 2 mg% was obtained.
失胤莢並
乳癌患者から採取した血清に、前記クロロホルムとメタ
ノールの混合液を用いた抽出を行う。Extraction is performed on serum collected from a breast cancer patient with lost seed capsules using the above-mentioned mixture of chloroform and methanol.
ｐＨ５，８で、メタノール溶液の上清と、硝酸銀の１．
０モル／Ｑ溶液とを１０：１の比率で混合して沈澱を生
成させ、沈澱中のシアル酸含量を測定する。 ２９．５
８■％なる値が得られた。At pH 5.8, supernatant of methanol solution and 1.
0 mol/Q solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured. 29.5
A value of 8% was obtained.
大凰孤並
乳癌患者から採取した血清に、前記クロロホルムとメタ
ノールの混合液を用いた抽出を行う。Serum collected from a patient with breast cancer is extracted using the mixture of chloroform and methanol.
ｐＨ１，５で、メタノール溶液の上滑と、塩化第二水銀
の１．０モルＩＱ溶液とを１０：１の比率で混合して沈
澱を生成させ、沈澱中のシアル酸含量を測定する。３３
．８９ｍｇ％なる値が得られた。At pH 1.5, a precipitate is formed by mixing a methanol solution and a 1.0 molar IQ solution of mercuric chloride in a ratio of 10:1, and the sialic acid content in the precipitate is measured. 33
．． A value of 89 mg% was obtained.
ス１１ルＵ
乳癌患者から採取した血清に、前記クロロホルムとメタ
ノールの混合液を用いた抽出を行う。Step 11 Extract the serum collected from a breast cancer patient using the mixture of chloroform and methanol.
ｐＨ３．６で、メタノール溶液の上清と、硫酸第二銅の
１．０モル／Ｑ溶液とをｌＯ：１の比率で混合して沈澱
を生成させ、沈澱中のシアル酸含量を測定する。At pH 3.6, the supernatant of the methanol solution and a 1.0 mol/Q solution of cupric sulfate are mixed at a ratio of 1O:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
３７．２０１１ｇ％なる値が得られた。A value of 37.2011 g% was obtained.
末嵐斑並
乳癌患者から採取した血清に、前記クロロホルムとメタ
ノールの混合液を用いた抽出を行う。Serum collected from a breast cancer patient with mild to moderate breast cancer is extracted using the above-mentioned mixture of chloroform and methanol.
ｐ）１５．３で、メタノール溶液の上清と、塩化カルシ
ウムの１．０モル／Ｑ溶液とを１０：１の比率で混合し
て沈澱を生成させ、沈澱中のシアル酸含量を測定する。p) At 15.3, the supernatant of the methanol solution and the 1.0 mol/Q solution of calcium chloride are mixed in a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
２８．１８■％なる値が得られた。A value of 28.18% was obtained.
大嵐豊川
乳癌患者から採取した血清に、前記クロロホルムとメタ
ノールの混合液を用いた抽出を行う。Serum collected from a Toyokawa breast cancer patient is extracted using the mixture of chloroform and methanol.
ｐ）１６．５で、メタノール溶液の上清と、塩化マグネ
シウムの１．０モルＩＱ溶液とを１０：１の比率で混合
して沈澱を生成させ、沈澱中のシアル酸含量を測定する
。２５．６７１１１ｇ％なる値が得られた。p) At 16.5, a precipitate is formed by mixing the supernatant of the methanol solution and a 1.0 molar IQ solution of magnesium chloride in a ratio of 10:1, and the sialic acid content in the precipitate is determined. A value of 25.67111 g% was obtained.
失胤涯赳
卵巣癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a deceased ovarian cancer patient is extracted using the mixture of chloroform and methanol.
ｐＨ５；３で、メタノール溶液の上清と、塩化ニッケル
の１．０モルＩＱ溶液とを１０＝１の比率で混合して沈
澱を生成させ、沈澱中のシアル酸含量を測定する。 ２
２．０９■％なる値が得られた。At pH 5:3, the supernatant of the methanol solution and a 1.0 molar IQ solution of nickel chloride are mixed in a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured. 2
A value of 2.09% was obtained.
失凰孤■
卵巣癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Loss of Death ■ Serum collected from an ovarian cancer patient is extracted using the above-mentioned mixture of chloroform and methanol.
ｐｉ１ｓ、３で、メタノール溶液の上清と、塩化カドミ
ウムの１．０モルＩＱ溶液とをｌＯ：１の比率で混合し
て沈澱を生成させ、沈澱中のシアル酸含量を測定する。In pils, 3, the supernatant of the methanol solution and a 1.0 molar IQ solution of cadmium chloride are mixed in a ratio of 1O:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
２５．８０ｍｇ％なる値が得られた。A value of 25.80 mg% was obtained.
失庭孤旦
卵巣癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with ovarian cancer is extracted using the mixture of chloroform and methanol.
ｐｉ（２，９で、メタノール溶液の上清と、硫酸第一鉄
の１．０モルＩＱ溶液とをｌＯ：１の比率で混合して沈
澱を生成させ、沈澱中のシアル酸含量を測定する。pi (2,9), mix the supernatant of the methanol solution and a 1.0 molar IQ solution of ferrous sulfate in a ratio of lO:1 to form a precipitate, and measure the sialic acid content in the precipitate. .
１８．７５．％なる値が得られた。18.75. A value of % was obtained.
失産貫旦
卵巣癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with ovarian cancer who has suffered from abortion is extracted using the above-mentioned mixture of chloroform and methanol.
ｐＨ１，７で、メタノール溶液の上清と、塩化亜鉛の１
．０モル／α溶液とを１０＝１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。At pH 1.7, the supernatant of methanol solution and 1 of zinc chloride
．． 0 mol/α solution at a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured.
２３．７７ｆｆＩｇ％なる値が得られた。A value of 23.77ffIg% was obtained.
失１貫旦
卵巣癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with ovarian cancer after one cancer is extracted using the above-mentioned mixture of chloroform and methanol.
ｐＨ１，７で、メタノール溶液の上清と、硫酸亜鉛の１
．０モル／Ｑ溶液とを１０：１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。At pH 1.7, the supernatant of methanol solution and 1 of zinc sulfate
．． 0 mol/Q solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
２６．８７■％なる値が得られた。A value of 26.87% was obtained.
尖凰盤Ｕ
卵巣癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Tsubouban U Serum collected from an ovarian cancer patient is extracted using the above-mentioned mixture of chloroform and methanol.
ｐ）１３．５で、メタノール溶液の上清と、硝酸鉛の１
．０モル／磨溶液とを１０：１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。p) At 13.5, the supernatant of the methanol solution and 1 of lead nitrate
．． 0 mol/polishing solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
２６．６３■％なる値が得られた。A value of 26.63% was obtained.
叉産涯鋲
卵巣癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from an ovarian cancer patient undergoing pregnancy is extracted using the mixture of chloroform and methanol.
ｐＨ５，８で、メタノール溶液の上清と、硝酸銀の１．
０モル／Ω溶液とをｌＯ：１の比率で混合して沈澱を生
成させ、沈澱中のシアル酸含量を測定する。At pH 5.8, supernatant of methanol solution and 1.
0 mol/Ω solution at a ratio of lO:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
２８．６６．１なる値が得られた。A value of 28.66.1 was obtained.
失凰五Ｕ
卵巣癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from an ovarian cancer patient is extracted using the mixture of chloroform and methanol.
ｐＨ１．５で、メタノール溶液の上清と、塩化第二水銀
の１．０モル／Ω溶液とを１０：１の比率で混合して沈
澱を生成させ、沈澱中のシアル酸含量を測定する。 ３
３．０８■％なる値が得られた。At pH 1.5, the supernatant of the methanol solution and a 1.0 mol/Ω solution of mercuric chloride are mixed at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured. 3
A value of 3.08% was obtained.
失産旌Ｕ
卵巣癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Abortion 旌U Serum collected from an ovarian cancer patient is extracted using the mixture of chloroform and methanol.
ｐＨ３．６で、メタノール溶液の上清と、硫酸第二銅の
１．０モル／Ω溶液とを１０：１の比率で混合して沈澱
を生成させ、沈澱中のシアル酸含量を測定する。At pH 3.6, the supernatant of the methanol solution and a 1.0 mol/Ω solution of cupric sulfate are mixed at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
２６．６３■％なる値が得られた。A value of 26.63% was obtained.
大適貫叩
卵巣癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with large-scale ovarian cancer is extracted using the mixture of chloroform and methanol.
ｐＨ５，３で、メタノール溶液の上清と、塩化カルシウ
ムの１．０モル／Ω溶液とを１０：１の比率で混合して
沈澱を生成させ、沈澱中のシアル酸含量を測定する。
１３．２６■％なる値が得られた。At pH 5.3, the supernatant of the methanol solution and a 1.0 mol/Ω solution of calcium chloride are mixed at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
A value of 13.26% was obtained.
大産五匹
卵巣癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Sera collected from five large-bred ovarian cancer patients are extracted using the above-mentioned mixture of chloroform and methanol.
ｐＨ６，５で、メタノール溶液の上清と、塩化マグネシ
ウムの１．０モル／Ω溶液とをｌＯ：１の比率で混合し
て沈澱を生成させ、沈澱中のシアル酸含量を測定する。At pH 6.5, the supernatant of the methanol solution and a 1.0 mol/Ω solution of magnesium chloride are mixed at a ratio of 1O:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
１１．８２ｍｇ％なる値が得られた。A value of 11.82 mg% was obtained.
失態■旦
頭部または頚部の癌患者から採取した血清に、前記クロ
ロホルムとメタノールの混合液を用いた抽出を行う、
ＰＨ５，３で、メタノール溶液の上清と。Blunder: Extracting serum collected from patients with cancer of the head or neck using the above-mentioned mixture of chloroform and methanol.
At pH 5.3, with the supernatant of the methanol solution.
塩化ニッケルの１．０モル／Ω溶液とを１０：１の比率
で混合して沈澱を生成させ、沈澱中のシアル酸含量を測
定する。 １２．６２■２なる値が得られた。A precipitate is formed by mixing with a 1.0 mol/Ω solution of nickel chloride at a ratio of 10:1, and the sialic acid content in the precipitate is measured. A value of 12.62■2 was obtained.
寒鳳五録 頭部または頚部の癌患者から採取した血清に。Kanho Goroku In serum collected from head or neck cancer patients.
前記クロロホルムとメタノールの混合液を用いた抽出を
行う、 ｐＨ５．３で、メタノール溶液の上清と、塩化
カドミウムの１．０モル／Ω溶液とをｌＯ：１の比率で
混合して沈澱を生成させ、沈澱中のシアル酸含量を測定
する。 ２１．９９■％なる値が得られた。Perform extraction using the mixture of chloroform and methanol. At pH 5.3, mix the supernatant of the methanol solution and a 1.0 mol/Ω solution of cadmium chloride at a ratio of 1O:1 to form a precipitate. and measure the sialic acid content in the precipitate. A value of 21.99% was obtained.
失凰叢閃
頭部または頚部の癌患者から採取した血清に、前記クロ
ロホルムとメタノールの混合液を用いた抽出を行う、
ｐｌ＋２．９で、メタノール溶液の上清と。Extracting the serum collected from a patient with cancer of the head or neck using the mixture of chloroform and methanol,
At pl+2.9, with the supernatant of the methanol solution.
硫酸第一鉄の１．０モル／Ω溶液とをｌＯ：１の比率で
混合して沈澱を生成させ、沈澱中のシアル酸含量を測定
する。 １６．０９■％なる値が得られた。A precipitate is formed by mixing with a 1.0 mol/Ω solution of ferrous sulfate at a ratio of 1O:1, and the sialic acid content in the precipitate is measured. A value of 16.09% was obtained.
失胤涯旦 頭部または頚部の癌患者から採取した血清に。Death of the Seed In serum collected from head or neck cancer patients.
前記クロロホルムとメタノールの混合液を用いた抽出を
行う、 ｐＨ１，７で、メタノール溶液の上清と、塩化
亜鉛の１．０モル／Ω溶液とを１０：１の比率で混合し
て沈澱を生成させ、沈澱中のシアル酸含量を測定する。Perform extraction using the mixture of chloroform and methanol. At pH 1.7, mix the supernatant of the methanol solution and a 1.0 mol/Ω solution of zinc chloride at a ratio of 10:1 to form a precipitate. and measure the sialic acid content in the precipitate.
１９．５６■％なる値が得られた。A value of 19.56% was obtained.
末五五並
頭部または頚部の癌患者から採取した血清に、前記クロ
ロホルムとメタノールの混合液を用いた抽出を行う、
ｐＨ１，７で、メタノール溶液の上清と、硫酸亜鉛の１
．０モル／Ω溶液とを１０：１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。２１．４
１■％なる値が得られた。Performing extraction using the mixture of chloroform and methanol on serum collected from a patient with cancer of the head or neck.
At pH 1.7, the supernatant of methanol solution and 1 of zinc sulfate
．． 0 mol/Ω solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured. 21.4
A value of 1% was obtained.
末流■並
頭部または頚部の癌患者から採取した血清に、前記クロ
ロホルムとメタノールの混合液を用いた抽出を行う、ｐ
Ｈ３．５で、メタノール溶液の上清と、硝酸鉛の１．０
モル／２溶液とを１０：１の比率で混合して沈澱を生成
させ、沈澱中のシアル酸含量を測定する。 ２９．９８
■％なる値が得られた。Endstream ■ Perform extraction using the above-mentioned mixture of chloroform and methanol on serum collected from patients with cancer of the head or neck, p
At H3.5, supernatant of methanol solution and 1.0% of lead nitrate
mol/2 solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured. 29.98
■A value of % was obtained.
ス】１町程 頭部または頚部の癌患者から採取した血清に。[S] About 1 town In serum collected from head or neck cancer patients.
前記クロロホルムとメタノールの混合液を用いた抽出を
行う、 ｐＨ５，８で、メタノール溶液の上溝と、硝酸
銀の１．０モルＩＱ溶液とを１０：１の比率で混合して
沈澱を生成させ、沈澱中のシアル酸含量を測定する。
１４．９３ｍｇ％なる値が得られた。Extraction is performed using a mixture of chloroform and methanol. At pH 5.8, the upper part of the methanol solution and a 1.0 molar IQ solution of silver nitrate are mixed in a ratio of 10:1 to form a precipitate. Measure the sialic acid content in the sample.
A value of 14.93 mg% was obtained.
失隨五閃 頭部または頚部の癌患者から採取した血清に。Lost Five Flashes In serum collected from head or neck cancer patients.
前記クロロホルムとメタノールの混合液を用い・た抽出
を行う、 ｐＨ１，５で、メタノール溶液の上清と、塩
化第二水銀の１．０モル／Ｑ溶液とを１０＝１の比率で
混合して沈澱を生成させ、沈澱中のシアル酸含量を測定
する。１８．８７．％なる値が得られた。Perform extraction using the mixture of chloroform and methanol, by mixing the supernatant of the methanol solution and a 1.0 mol/Q solution of mercuric chloride at a ratio of 10=1 at pH 1.5. A precipitate is generated and the sialic acid content in the precipitate is measured. 18.87. A value of % was obtained.
叉凰且利
頭部または頚部の癌患者から採取した血清に、前記クロ
ロホルムとメタノールの混合液を用いた抽出を行う、
ｐＨ３．６で、メタノール溶液の上清と。Extracting serum collected from a head or neck cancer patient using the mixture of chloroform and methanol;
with the supernatant of the methanol solution at pH 3.6.
硫酸第二銅の１．０モルＩＱ溶液とを１０：１の比率で
混合して沈澱を生成させ、沈澱中のシアル酸含量を測定
する。 １４．７０■％なる値が得られた。A precipitate is formed by mixing with a 1.0 molar IQ solution of cupric sulfate in a ratio of 10:1, and the sialic acid content in the precipitate is measured. A value of 14.70% was obtained.
失直涯銭
頭部または頚部の癌患者から採取した血清に、前記クロ
ロホルムとメタノールの混合液を用いた抽出を行う、
ｐＨ５，３で、メタノール溶液の上清と。Performing extraction using the mixture of chloroform and methanol on serum collected from a paralyzed or cervical cancer patient;
With the supernatant of methanol solution at pH 5.3.
塩化カルシウムの１．０モルＩＱ溶液とを１０＝１の比
率で混合して沈澱を生成させ、沈澱中のシアル酸含量を
測定する。 ９．７２［％なる値が得られた。A precipitate is formed by mixing with a 1.0 molar IQ solution of calcium chloride in a ratio of 10=1, and the sialic acid content in the precipitate is measured. A value of 9.72% was obtained.
叉適叢虹
頭部または頚部の癌患者から採取した血清に、前記クロ
ロホルムとメタノールの混合液を用いた抽出を行う、
ｐｌ（６，５で、メタノール溶液の上清と、塩化マグネ
シウムの１．０モル／Ｑ溶液とをＬＯ：１の比率で混合
して沈澱を生成させ、沈澱中のシアル酸含量を測定する
。 １２．２７Ｑ％なる値が得られた。Performing extraction using the mixture of chloroform and methanol on the serum collected from a patient with cancer of the head or neck.
pl (6,5), the supernatant of the methanol solution and a 1.0 mol/Q solution of magnesium chloride are mixed at a ratio of LO:1 to form a precipitate, and the sialic acid content in the precipitate is measured. A value of 12.27Q% was obtained.
来産叢且
腺癌の患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with paroxysmal adenocarcinoma is extracted using the mixture of chloroform and methanol.
ｐＨ５，３で、メタノール溶液の上清と、塩化ニッケル
の１．Ｏモル／β溶液とを１０：１の比率で混合して沈
澱を生成させ、沈澱中のシアル酸含量を測定する。 １
１．７７ｍｇ％なる値が得られた。At pH 5.3, supernatant of methanol solution and nickel chloride 1. O mol/β solution is mixed at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured. 1
A value of 1.77 mg% was obtained.
叉五且邸
腺癌の患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with pharyngeal adenocarcinoma is extracted using the mixture of chloroform and methanol.
ＰＨ５，３で、メタノール溶液の上清と、塩化カドミウ
ムの１．０モル／Ｑ溶液とを１０：１の比率で混合して
沈澱を生成させ、沈澱中のシアル酸含量を測定する。
２０．０９■％なる値が得られた。At pH 5.3, the supernatant of the methanol solution and a 1.0 mol/Q solution of cadmium chloride are mixed at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
A value of 20.09% was obtained.
失凰孤旦
腺癌の患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with phlegmon adenocarcinoma is extracted using the mixture of chloroform and methanol.
ｐＨ２，９で、メタノール溶液の上清と、硫酸第一鉄の
１．０モル／Ｑ溶液とを１０＝１の比率で混合して沈澱
を生成させ、沈澱中のシアル酸含量を測定する。At pH 2.9, the supernatant of the methanol solution and a 1.0 mol/Q solution of ferrous sulfate are mixed at a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured.
１６．１３■％なる値が得られた。A value of 16.13% was obtained.
大適貫並
腺癌の患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Extraction is performed on serum collected from a patient with large-scale transverse adenocarcinoma using the above-mentioned mixture of chloroform and methanol.
ｐＨ１．７で、メタノール溶液の上清と、塩化亜鉛の１
．０モル／ｌ溶液とを１０：１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。At pH 1.7, the supernatant of the methanol solution and 1
．． 0 mol/l solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
１８．６９■ヌなる値が得られた。A value of 18.69μ was obtained.
去１目シ獲
腺癌の患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with thyroid carcinoma is subjected to extraction using the above-mentioned mixture of chloroform and methanol.
ｐＨ１，７で、メタノール溶液の上清と、硫酸亜鉛の１
．０モルＩＱ溶液とを１０：１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。At pH 1.7, the supernatant of methanol solution and 1 of zinc sulfate
．． A precipitate is formed by mixing with a 0 molar IQ solution in a ratio of 10:1, and the sialic acid content in the precipitate is measured.
１７．１５■％なる値が得られた。A value of 17.15% was obtained.
叉凰貫虹
腺癌の患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Extraction is performed on serum collected from a patient with diadenoid carcinoma using the above-mentioned mixture of chloroform and methanol.
ｐＨ３，５で、メタノール溶液の上清と、硝酸鉛の１．
０モル／Ｑ溶液とを１０：１の比率で混合して沈澱を生
成させ、沈澱中のシアル酸含量を測定する。At pH 3.5, supernatant of methanol solution and 1.0% of lead nitrate.
0 mol/Q solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
２２．６６■算なる値が得られた。A value of 22.66 mm was obtained.
太産涯並
腺癌の患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Extraction is performed on serum collected from a patient with adenocarcinoma of the infant stage using the above-mentioned mixture of chloroform and methanol.
ＰＨ５，８で、メタノール溶液の上清と、硝酸銀の１．
０モル／党溶液とを１０：１の比率で混合して沈澱を生
成させ、沈澱中のシアル酸含量を測定する。At pH 5.8, supernatant of methanol solution and silver nitrate 1.
A precipitate is formed by mixing 0 mol/part solution at a ratio of 10:1, and the sialic acid content in the precipitate is measured.
１６．５１■％なる値が得られた。A value of 16.51% was obtained.
失五叢並
腺癌の患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with aplasic adenocarcinoma is extracted using the mixture of chloroform and methanol.
ｐＨ１．５で、メタノール溶液の上清と、塩化第二水銀
の１．０モルＩＱ溶液とを１０：１の比率で混合して沈
澱を生成させ、沈澱中のシアル酸含量を測定する。１８
．９４■ぶなる値が得られた。At pH 1.5, the supernatant of the methanol solution and a 1.0 molar IQ solution of mercuric chloride are mixed in a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured. 18
．． A value of 94 ■ was obtained.
失履■到
腺癌の患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with lapsed adenocarcinoma is extracted using the mixture of chloroform and methanol.
ｐＨ３．６で、メタノール溶液の上清と、硫酸第二銅の
１．０モル／Ｑ溶液とを１０：１の比率で混合して沈澱
を生成させ、沈澱中のシアル酸含量を測定する。At pH 3.6, the supernatant of the methanol solution and a 1.0 mol/Q solution of cupric sulfate are mixed at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
１７．４１■％なる値が得られた。A value of 17.41% was obtained.
叉胤−旦
腺癌の患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with cancerous adenocarcinoma is extracted using the mixture of chloroform and methanol.
ＰＨ５，３で、メタノール溶液の上清と、塩化カルシウ
ムの１．０モルＩＱ溶液とをｌＯ：１の比率で混合して
沈澱を生成させ、沈澱中のシアル酸含量を測定する。
９．７３■でなる値が得られた。At pH 5.3, the supernatant of the methanol solution and a 1.0 molar IQ solution of calcium chloride are mixed in a ratio of 1O:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
A value of 9.73■ was obtained.
寒直且Ｕ
腺癌の患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with cold adenocarcinoma is extracted using the mixture of chloroform and methanol.
ｐＨ６，５で、メタノール溶液の上清と、塩化マグネシ
ウムの１．０モル／１１溶液とを１０＝１の比率で混合
して沈澱を生成させ、沈澱中のシアル酸含量を測定する
。 ７．９４ｍｇ％なる値が得られた。At pH 6.5, the supernatant of the methanol solution and a 1.0 mol/11 solution of magnesium chloride are mixed in a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured. A value of 7.94 mg% was obtained.
大嵐五互
結腸癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with Oarashi Goko colon cancer is extracted using the above-mentioned mixture of chloroform and methanol.
ｐ＋＋ｓ、３で、メタノール溶液の上清と、塩化ニッケ
ルの１．０モルＩＱ溶液とを１０＝１の比率で混合して
沈澱を生成させ、沈澱中のシアル酸含量を測定する。
２０．７０■％なる値が得られた。At p++s, 3, the supernatant of the methanol solution and a 1.0 molar IQ solution of nickel chloride are mixed in a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured.
A value of 20.70% was obtained.
笑適叢り
結腸癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a colon cancer patient is extracted using the mixture of chloroform and methanol.
ＰＨ５，３で、メタノール溶液の上清と、塩化カドミウ
ムの１．０モル／Ｑ溶液とを１０：１の比率で混合して
沈澱を生成させ、沈澱中のシアル酸含量を測定する。
３２．１４■％なる値が得られた。At pH 5.3, the supernatant of the methanol solution and a 1.0 mol/Q solution of cadmium chloride are mixed at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
A value of 32.14% was obtained.
叉産■互
結腸癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with reciprocal colon cancer is extracted using the mixture of chloroform and methanol.
ＰＨ２，９で、メタノール溶液の上清と、硫酸第一鉄の
１．０モル／１２溶液とを１０＝１の比率で混合して沈
澱を生成させ、沈澱中のシアル酸含量を測定する。At pH 2.9, the supernatant of the methanol solution and a 1.0 mol/12 solution of ferrous sulfate are mixed in a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured.
２４．２３■％なる値が得られた。A value of 24.23% was obtained.
叉産叢匹
結腸癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a colon cancer patient with multiple births is extracted using the above-mentioned mixture of chloroform and methanol.
ＰＩ（１，７で、メタノール溶液の上清と、塩化亜鉛の
１．０モル／Ｑ溶液とを１０＝１の比率で混合して沈澱
を生成させ、沈澱中のシアル酸含量を測定する。PI (1,7), the supernatant of the methanol solution and the 1.0 mol/Q solution of zinc chloride are mixed in a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured.
２７．５２■石なる値が得られた。A value of 27.52 stone was obtained.
スＪＪＩＺＺ
結腸癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。SJJIZZ Serum collected from a colon cancer patient is extracted using the mixture of chloroform and methanol.
ｐＨ１．７で、メタノール溶液の上清と、硫酸亜鉛の１
．０モル／Ｑ溶液とを１０：１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。At pH 1.7, the supernatant of the methanol solution and the solution of zinc sulfate
．． 0 mol/Q solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
３２．６３■メなる値が得られた。A value of 32.63 mm was obtained.
大嵐叢亙
結腸癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from patients with colon cancer in the Great Arashi region is extracted using the above-mentioned mixture of chloroform and methanol.
ｐ）１３．５で、メタノール溶液の上清と、硝酸鉛の１
．０モル／ρ溶液とを１０：１の比率で混合して沈澱を
生成させ、沈澱中のシアル酸含量を測定する。p) At 13.5, the supernatant of the methanol solution and 1 of lead nitrate
．． 0 mol/ρ solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
３５．５５ｍｇ％なる値が得られた。A value of 35.55 mg% was obtained.
末直！７ｕ
結腸癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Sue Nao! 7u Extraction is performed on serum collected from a colon cancer patient using the above-mentioned mixture of chloroform and methanol.
ｐＨ５，８で、メタノール溶液の上清と、硝酸銀の１．
０モル／Ｑ溶液とを１０：１の比率で混合して沈澱を生
成させ、沈澱中のシアル酸含量を測定する。At pH 5.8, supernatant of methanol solution and 1.
0 mol/Q solution at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
２９．４６Ｎ％なる値が得られた。A value of 29.46N% was obtained.
塞直銖級
結腸癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with colon cancer is extracted using the mixture of chloroform and methanol.
ｐ！（１，５で、メタノール溶液の上清と、塩化第二水
銀の１．０モルＩＱ溶液とを１０＝１の比率で混合して
沈澱を生成させ、沈澱中のシアル酸含量を測定する。
３１．６６■算なる値が得られた。p! (In steps 1 and 5, the supernatant of the methanol solution and the 1.0 molar IQ solution of mercuric chloride are mixed in a ratio of 10=1 to form a precipitate, and the sialic acid content in the precipitate is measured.
A value of 31.66 mm was obtained.
末適透旦
結腸癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a colon cancer patient is extracted using the mixture of chloroform and methanol.
ｐＨ３．６で、メタノール溶液の上滑と、硫酸第二銅の
１．０モル／ａ溶液とを１０：１の比率で混合して沈澱
を生成させ、沈澱中のシアル酸含量を測定する。At pH 3.6, a methanol solution and a 1.0 mol/a solution of cupric sulfate are mixed in a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
３２．５１■％なる値が得られた。A value of 32.51% was obtained.
叉凰■閃
結腸癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Serum collected from a patient with colon cancer is extracted using the mixture of chloroform and methanol.
ｐＨ５，３で５メタノール溶液の上滑と、塩化カルシウ
ムの１．０モルＩＱ溶液とをｌＯ：１の比率で混合して
沈澱を生成させ、沈澱中のシアル酸含量を測定する。
Ｌ８．５１ｍｇ％なる値が得られた。At pH 5.3, a precipitate is formed by mixing a 1.0 molar IQ solution of calcium chloride with a 1.0 molar IQ solution of calcium chloride at pH 5.3, and the sialic acid content in the precipitate is measured.
A value of 8.51 mg% L was obtained.
失凰鼠Ｕ
結腸癌患者から採取した血清に、前記クロロホルムとメ
タノールの混合液を用いた抽出を行う。Loss of Phlegm U Serum collected from a colon cancer patient is extracted using the above-mentioned mixture of chloroform and methanol.
ｐＨ６，５で、メタノール溶液の上清と、塩化マグネシ
ウムの１．０モル／Ｑ溶液とを１０：１の比率で混合し
て沈澱を生成させ、沈澱中のシアル酸含量を測定する。At pH 6.5, the supernatant of the methanol solution and a 1.0 mol/Q solution of magnesium chloride are mixed at a ratio of 10:1 to form a precipitate, and the sialic acid content in the precipitate is measured.
１６．６ＪＩ■２なる値が得られた。A value of 16.6JI■2 was obtained.
Claims (12)
溶媒または溶媒混合液を用いた抽出によって中性脂質を
除去し、その水相から、シアル酸を有する糖脂質を含有
するリポ蛋白質分画を沈澱させ、沈澱した分画を再溶解
させ、溶液のシアル酸含有量、あるいはシアル酸を有す
る糖脂質の量および質を、公知の方法を用いて測定する
ことからなるシアル酸を有する血中糖脂質複合体の選択
的測定を行なうための分析方法であって、 沈澱剤として、混合液中の最終金属イオン濃度が０．０
０５〜０．１モル／ｌに対応する量の、アルカリ金属以
外の１価、２価、あるいは３価の水溶性金属塩を加え、
かつ、ｐＨ１．５〜８．０にて沈澱を生成させることを
特徴とする、シアル酸を有する血中糖脂質複合体の選択
的測定を行なうための分析方法。(1) Serum is diluted with water, neutral lipids are removed by extraction with a water-immiscible organic solvent or solvent mixture, and lipoproteins containing glycolipids with sialic acid are extracted from the aqueous phase. having sialic acid, comprising precipitating a fraction, redissolving the precipitated fraction, and measuring the sialic acid content of the solution, or the quantity and quality of glycolipids having sialic acid, using known methods. An analytical method for selectively measuring blood glycolipid complexes, the method comprising using a precipitant as a precipitant with a final metal ion concentration of 0.0 in the mixed solution.
Adding a monovalent, divalent, or trivalent water-soluble metal salt other than an alkali metal in an amount corresponding to 0.05 to 0.1 mol/l,
An analytical method for selectively measuring a blood glycolipid complex having sialic acid, the method comprising: producing a precipitate at pH 1.5 to 8.0.
＋）の塩を加え、かつｐＨ３．０〜３．５にて沈澱を生
成させることを特徴とする請求項（１）記載の分析方法
。(2) Water-soluble ferrous ion (Fe^2^
2. The analytical method according to claim 1, wherein a salt of (+) is added and a precipitate is formed at a pH of 3.0 to 3.5.
＋）の塩を加え、かつｐＨ３．０〜５．０にて沈澱を生
成させることを特徴とする請求項（１）記載の分析方法
。(3) Water-soluble ferric ion (Fe^3^
The method of analysis according to claim 1, characterized in that a salt of (+) is added and a precipitate is formed at a pH of 3.0 to 5.0.
＾＋）の塩を加え、かつｐＨ４．０〜５．５にて沈澱を
生成させることを特徴とする請求項（１）記載の分析方
法。(4) Water-soluble cobalt ion (Co^2
The analytical method according to claim 1, characterized in that a salt of ^+) is added and a precipitate is formed at pH 4.0 to 5.5.
＾＋）の塩を加え、かつｐＨ４．５〜５．５にて沈澱を
生成させることを特徴とする請求項（１）記載の分析方
法。(5) As a precipitant, water-soluble nickel ions (Ni^2
The analytical method according to claim 1, characterized in that a salt of ^+) is added and a precipitate is formed at a pH of 4.5 to 5.5.
２＾＋）の塩を加え、かつｐＨ４．５〜６．０にて沈澱
を生成させることを特徴とする請求項（１）記載の分析
方法。(6) Water-soluble cadmium ion (Cd^
2^+) salt is added and a precipitate is formed at a pH of 4.5 to 6.0.
）の塩を加え、かつｐＨ１．５〜５．０にて沈澱を生成
させることを特徴とする請求項（１）記載の分析方法。(7) Water-soluble zinc ion (Zn^2^+
The analytical method according to claim 1, characterized in that a salt of ) is added and a precipitate is formed at a pH of 1.5 to 5.0.
を加え、かつｐＨ６．０〜８．０にて沈澱を生成させる
ことを特徴とする請求項（１）記載の分析方法。(8) The analytical method according to claim (1), characterized in that a water-soluble silver ion (Ag^+) salt is added as a precipitant, and the precipitate is formed at a pH of 6.0 to 8.0.
＾＋）の塩を加え、かつｐＨ１．５〜３．０にて沈澱を
生成させることを特徴とする請求項（１）記載の分析方
法。(9) Water-soluble mercury ion (Hg^2
The analytical method according to claim 1, characterized in that a salt of ^+) is added and a precipitate is formed at a pH of 1.5 to 3.0.
＾＋）の塩を加え、かつｐＨ３．０〜４．５にて沈澱を
生成させることを特徴とする請求項（１）記載の分析方
法。(10) Water-soluble cupric ion (Cu^2
The analytical method according to claim 1, characterized in that a salt of ^+) is added and a precipitate is formed at a pH of 3.0 to 4.5.
＾２＾＋）の塩を加え、かつｐＨ４．５〜５．５にて沈
澱を生成させることを特徴とする請求項（１）記載の分
析方法。(11) Water-soluble calcium ions (Ca
The analytical method according to claim 1, characterized in that a salt of ^2^+) is added and a precipitate is formed at a pH of 4.5 to 5.5.
ｇ＾２＾＋）の塩を加え、かつｐＨ５．５〜７．０にて
沈澱を生成させることを特徴とする請求項（１）記載の
分析方法。(12) Water-soluble magnesium ion (M
The analytical method according to claim 1, characterized in that a salt of g^2^+) is added and a precipitate is formed at a pH of 5.5 to 7.0.
Applications Claiming Priority (4)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| HU24989 | 1989-01-23 | ||
| HU2251-249/89 | 1989-01-23 | ||
| HU2251-2190/89 | 1989-05-08 | ||
| HU219089A HU208377B (en) | 1989-05-08 | 1989-05-08 | Analytic method for selective detecting glycolipid complexes carrying sial acid |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH02206763A true JPH02206763A (en) | 1990-08-16 |
Family
ID=26317184
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP1131718A Pending JPH02206763A (en) | 1989-01-23 | 1989-05-26 | Analysis for selective measurement of glycolipid composite in blood having sialic acid |
Country Status (10)
| Country | Link |
|---|---|
| EP (1) | EP0515355A1 (en) |
| JP (1) | JPH02206763A (en) |
| KR (1) | KR910700457A (en) |
| CN (1) | CN1044537A (en) |
| AU (1) | AU3735689A (en) |
| DK (1) | DK226590D0 (en) |
| ES (1) | ES2012728A6 (en) |
| FI (1) | FI913518A0 (en) |
| NO (1) | NO903877D0 (en) |
| WO (1) | WO1990008317A1 (en) |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4342567A (en) * | 1981-07-06 | 1982-08-03 | Sloan Kettering Institute For Cancer Research | Sialic acid determination method |
-
1989
- 1989-05-12 EP EP89905744A patent/EP0515355A1/en not_active Ceased
- 1989-05-12 WO PCT/HU1989/000021 patent/WO1990008317A1/en not_active Application Discontinuation
- 1989-05-12 KR KR1019900701970A patent/KR910700457A/en not_active Application Discontinuation
- 1989-05-12 AU AU37356/89A patent/AU3735689A/en not_active Abandoned
- 1989-05-26 JP JP1131718A patent/JPH02206763A/en active Pending
- 1989-06-06 ES ES8901972A patent/ES2012728A6/en not_active Expired - Fee Related
- 1989-06-20 CN CN89104207A patent/CN1044537A/en active Pending
-
1990
- 1990-09-05 NO NO1990903877A patent/NO903877D0/en unknown
- 1990-09-20 DK DK226590A patent/DK226590D0/en not_active Application Discontinuation
-
1991
- 1991-07-23 FI FI913518A patent/FI913518A0/en not_active Application Discontinuation
Also Published As
| Publication number | Publication date |
|---|---|
| WO1990008317A1 (en) | 1990-07-26 |
| FI913518A0 (en) | 1991-07-23 |
| KR910700457A (en) | 1991-03-15 |
| CN1044537A (en) | 1990-08-08 |
| ES2012728A6 (en) | 1990-04-01 |
| AU3735689A (en) | 1990-08-13 |
| DK226590A (en) | 1990-09-20 |
| DK226590D0 (en) | 1990-09-20 |
| EP0515355A1 (en) | 1992-12-02 |
| NO903877L (en) | 1990-09-05 |
| NO903877D0 (en) | 1990-09-05 |
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