CN112630216A - 一种检测1,5-ag的试剂盒和检测方法 - Google Patents
一种检测1,5-ag的试剂盒和检测方法 Download PDFInfo
- Publication number
- CN112630216A CN112630216A CN202011363454.7A CN202011363454A CN112630216A CN 112630216 A CN112630216 A CN 112630216A CN 202011363454 A CN202011363454 A CN 202011363454A CN 112630216 A CN112630216 A CN 112630216A
- Authority
- CN
- China
- Prior art keywords
- reagent
- sample
- kit
- oxidase
- standard
- 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
- MPCAJMNYNOGXPB-SLPGGIOYSA-N 1,5-anhydro-D-glucitol Chemical compound OC[C@H]1OC[C@H](O)[C@@H](O)[C@@H]1O MPCAJMNYNOGXPB-SLPGGIOYSA-N 0.000 title claims abstract description 38
- 238000000034 method Methods 0.000 title abstract description 14
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract description 47
- 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 claims abstract description 14
- 239000008103 glucose Substances 0.000 claims abstract description 12
- 239000003755 preservative agent Substances 0.000 claims abstract description 12
- 230000002335 preservative effect Effects 0.000 claims abstract description 12
- 239000003381 stabilizer Substances 0.000 claims abstract description 12
- 238000001514 detection method Methods 0.000 claims abstract description 11
- 239000006174 pH buffer Substances 0.000 claims abstract description 10
- 108010001816 pyranose oxidase Proteins 0.000 claims abstract description 10
- 239000004094 surface-active agent Substances 0.000 claims abstract description 10
- 108010015428 Bilirubin oxidase Proteins 0.000 claims abstract description 7
- 108010015776 Glucose oxidase Proteins 0.000 claims abstract description 7
- 239000004366 Glucose oxidase Substances 0.000 claims abstract description 7
- 102000003992 Peroxidases Human genes 0.000 claims abstract description 7
- 229940116332 glucose oxidase Drugs 0.000 claims abstract description 7
- 235000019420 glucose oxidase Nutrition 0.000 claims abstract description 7
- 108040007629 peroxidase activity proteins Proteins 0.000 claims abstract description 7
- -1 saccharide compounds Chemical class 0.000 claims abstract description 7
- RLFWWDJHLFCNIJ-UHFFFAOYSA-N 4-aminoantipyrine Chemical compound CN1C(C)=C(N)C(=O)N1C1=CC=CC=C1 RLFWWDJHLFCNIJ-UHFFFAOYSA-N 0.000 claims abstract description 6
- 102000004190 Enzymes Human genes 0.000 claims abstract description 6
- 108090000790 Enzymes Proteins 0.000 claims abstract description 6
- 230000003281 allosteric effect Effects 0.000 claims abstract description 6
- 229940088598 enzyme Drugs 0.000 claims abstract description 6
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Natural products OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 claims abstract description 4
- IRQRBVOQGUPTLG-UHFFFAOYSA-M sodium;3-(n-ethyl-3-methylanilino)-2-hydroxypropane-1-sulfonate Chemical compound [Na+].[O-]S(=O)(=O)CC(O)CN(CC)C1=CC=CC(C)=C1 IRQRBVOQGUPTLG-UHFFFAOYSA-M 0.000 claims abstract description 4
- 235000010323 ascorbic acid Nutrition 0.000 claims abstract description 3
- 239000011668 ascorbic acid Substances 0.000 claims abstract description 3
- 108090000854 Oxidoreductases Proteins 0.000 claims abstract 2
- 102000004316 Oxidoreductases Human genes 0.000 claims abstract 2
- 229960005070 ascorbic acid Drugs 0.000 claims abstract 2
- 229940072417 peroxidase Drugs 0.000 claims abstract 2
- 238000002835 absorbance Methods 0.000 claims description 12
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N Sodium azide Chemical compound [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 claims description 8
- 239000007853 buffer solution Substances 0.000 claims description 6
- 238000006243 chemical reaction Methods 0.000 claims description 5
- 239000000126 substance Substances 0.000 claims description 5
- 108010024957 Ascorbate Oxidase Proteins 0.000 claims description 4
- 238000003556 assay Methods 0.000 claims description 4
- GPRLSGONYQIRFK-MNYXATJNSA-N triton Chemical compound [3H+] GPRLSGONYQIRFK-MNYXATJNSA-N 0.000 claims description 4
- 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 claims 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 claims description 2
- CHHHXKFHOYLYRE-UHFFFAOYSA-M 2,4-Hexadienoic acid, potassium salt (1:1), (2E,4E)- Chemical compound [K+].CC=CC=CC([O-])=O CHHHXKFHOYLYRE-UHFFFAOYSA-M 0.000 claims 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 claims description 2
- DVLFYONBTKHTER-UHFFFAOYSA-N 3-(N-morpholino)propanesulfonic acid Chemical compound OS(=O)(=O)CCCN1CCOCC1 DVLFYONBTKHTER-UHFFFAOYSA-N 0.000 claims description 2
- NUFBIAUZAMHTSP-UHFFFAOYSA-N 3-(n-morpholino)-2-hydroxypropanesulfonic acid Chemical compound OS(=O)(=O)CC(O)CN1CCOCC1 NUFBIAUZAMHTSP-UHFFFAOYSA-N 0.000 claims description 2
- 239000007988 ADA buffer Substances 0.000 claims description 2
- CEAZRRDELHUEMR-URQXQFDESA-N Gentamicin Chemical compound O1[C@H](C(C)NC)CC[C@@H](N)[C@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](NC)[C@@](C)(O)CO2)O)[C@H](N)C[C@@H]1N CEAZRRDELHUEMR-URQXQFDESA-N 0.000 claims description 2
- 229930182566 Gentamicin Natural products 0.000 claims description 2
- 239000007995 HEPES buffer Substances 0.000 claims description 2
- 239000007993 MOPS buffer Substances 0.000 claims description 2
- JOCBASBOOFNAJA-UHFFFAOYSA-N N-tris(hydroxymethyl)methyl-2-aminoethanesulfonic acid Chemical compound OCC(CO)(CO)NCCS(O)(=O)=O JOCBASBOOFNAJA-UHFFFAOYSA-N 0.000 claims description 2
- 229930193140 Neomycin Natural products 0.000 claims description 2
- 239000007990 PIPES buffer Substances 0.000 claims description 2
- 229910019142 PO4 Inorganic materials 0.000 claims description 2
- 239000007994 TES buffer Substances 0.000 claims description 2
- 239000007983 Tris buffer Substances 0.000 claims description 2
- OWMVSZAMULFTJU-UHFFFAOYSA-N bis-tris Chemical compound OCCN(CCO)C(CO)(CO)CO OWMVSZAMULFTJU-UHFFFAOYSA-N 0.000 claims description 2
- 239000008139 complexing agent Substances 0.000 claims description 2
- 229960002518 gentamicin Drugs 0.000 claims description 2
- 229910021645 metal ion Inorganic materials 0.000 claims description 2
- 229960004927 neomycin Drugs 0.000 claims description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 claims description 2
- 239000010452 phosphate Substances 0.000 claims description 2
- 229920001223 polyethylene glycol Polymers 0.000 claims description 2
- 229920000136 polysorbate Polymers 0.000 claims description 2
- 239000004302 potassium sorbate Substances 0.000 claims description 2
- 235000010241 potassium sorbate Nutrition 0.000 claims description 2
- 229940069338 potassium sorbate Drugs 0.000 claims description 2
- 150000005846 sugar alcohols Polymers 0.000 claims description 2
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 claims description 2
- QTBSBXVTEAMEQO-UHFFFAOYSA-M Acetate Chemical compound CC([O-])=O QTBSBXVTEAMEQO-UHFFFAOYSA-M 0.000 claims 1
- BTBUEUYNUDRHOZ-UHFFFAOYSA-N Borate Chemical compound [O-]B([O-])[O-] BTBUEUYNUDRHOZ-UHFFFAOYSA-N 0.000 claims 1
- BVKZGUZCCUSVTD-UHFFFAOYSA-L Carbonate Chemical compound [O-]C([O-])=O BVKZGUZCCUSVTD-UHFFFAOYSA-L 0.000 claims 1
- KRKNYBCHXYNGOX-UHFFFAOYSA-K Citrate Chemical compound [O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O KRKNYBCHXYNGOX-UHFFFAOYSA-K 0.000 claims 1
- 210000002966 serum Anatomy 0.000 abstract description 17
- 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 abstract description 11
- 239000003795 chemical substances by application Substances 0.000 abstract description 4
- WQZGKKKJIJFFOK-QTVWNMPRSA-N D-mannopyranose Chemical compound OC[C@H]1OC(O)[C@@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-QTVWNMPRSA-N 0.000 abstract description 2
- 229930091371 Fructose Natural products 0.000 abstract description 2
- 239000005715 Fructose Substances 0.000 abstract description 2
- RFSUNEUAIZKAJO-ARQDHWQXSA-N Fructose Chemical compound OC[C@H]1O[C@](O)(CO)[C@@H](O)[C@@H]1O RFSUNEUAIZKAJO-ARQDHWQXSA-N 0.000 abstract description 2
- 230000008569 process Effects 0.000 abstract description 2
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 10
- 230000000694 effects Effects 0.000 description 6
- 238000011084 recovery Methods 0.000 description 5
- 238000012360 testing method Methods 0.000 description 5
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 4
- 206010012601 diabetes mellitus Diseases 0.000 description 4
- 239000012086 standard solution Substances 0.000 description 4
- 210000002700 urine Anatomy 0.000 description 4
- ZKHQWZAMYRWXGA-KQYNXXCUSA-J ATP(4-) Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](COP([O-])(=O)OP([O-])(=O)OP([O-])([O-])=O)[C@@H](O)[C@H]1O ZKHQWZAMYRWXGA-KQYNXXCUSA-J 0.000 description 3
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- ZKHQWZAMYRWXGA-UHFFFAOYSA-N Adenosine triphosphate Natural products C1=NC=2C(N)=NC=NC=2N1C1OC(COP(O)(=O)OP(O)(=O)OP(O)(O)=O)C(O)C1O ZKHQWZAMYRWXGA-UHFFFAOYSA-N 0.000 description 3
- XJLXINKUBYWONI-DQQFMEOOSA-N [[(2r,3r,4r,5r)-5-(6-aminopurin-9-yl)-3-hydroxy-4-phosphonooxyoxolan-2-yl]methoxy-hydroxyphosphoryl] [(2s,3r,4s,5s)-5-(3-carbamoylpyridin-1-ium-1-yl)-3,4-dihydroxyoxolan-2-yl]methyl phosphate Chemical compound NC(=O)C1=CC=C[N+]([C@@H]2[C@H]([C@@H](O)[C@H](COP([O-])(=O)OP(O)(=O)OC[C@@H]3[C@H]([C@@H](OP(O)(O)=O)[C@@H](O3)N3C4=NC=NC(N)=C4N=C3)O)O2)O)=C1 XJLXINKUBYWONI-DQQFMEOOSA-N 0.000 description 3
- 230000009471 action Effects 0.000 description 3
- 229910052799 carbon Inorganic materials 0.000 description 3
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 3
- 238000005259 measurement Methods 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- BPYKTIZUTYGOLE-IFADSCNNSA-N Bilirubin Chemical compound N1C(=O)C(C)=C(C=C)\C1=C\C1=C(C)C(CCC(O)=O)=C(CC2=C(C(C)=C(\C=C/3C(=C(C=C)C(=O)N\3)C)N2)CCC(O)=O)N1 BPYKTIZUTYGOLE-IFADSCNNSA-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
- 102000030595 Glucokinase Human genes 0.000 description 2
- 108010021582 Glucokinase Proteins 0.000 description 2
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical group [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 230000018109 developmental process Effects 0.000 description 2
- 238000003745 diagnosis Methods 0.000 description 2
- 229930027945 nicotinamide-adenine dinucleotide Natural products 0.000 description 2
- 230000009103 reabsorption Effects 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 230000008929 regeneration Effects 0.000 description 2
- 238000011069 regeneration method Methods 0.000 description 2
- 238000012827 research and development Methods 0.000 description 2
- VRYALKFFQXWPIH-PBXRRBTRSA-N (3r,4s,5r)-3,4,5,6-tetrahydroxyhexanal Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)CC=O VRYALKFFQXWPIH-PBXRRBTRSA-N 0.000 description 1
- MPCAJMNYNOGXPB-UHFFFAOYSA-N 1,5-Anhydro-mannit Natural products OCC1OCC(O)C(O)C1O MPCAJMNYNOGXPB-UHFFFAOYSA-N 0.000 description 1
- OCLOLUFOLJIQDC-HSUXUTPPSA-N 1,5-anhydro-D-fructose Chemical compound OC[C@H]1OCC(=O)[C@@H](O)[C@@H]1O OCLOLUFOLJIQDC-HSUXUTPPSA-N 0.000 description 1
- 108060006004 Ascorbate peroxidase Proteins 0.000 description 1
- NBSCHQHZLSJFNQ-GASJEMHNSA-N D-Glucose 6-phosphate Chemical compound OC1O[C@H](COP(O)(O)=O)[C@@H](O)[C@H](O)[C@H]1O NBSCHQHZLSJFNQ-GASJEMHNSA-N 0.000 description 1
- PHOQVHQSTUBQQK-SQOUGZDYSA-N D-glucono-1,5-lactone Chemical compound OC[C@H]1OC(=O)[C@H](O)[C@@H](O)[C@@H]1O PHOQVHQSTUBQQK-SQOUGZDYSA-N 0.000 description 1
- VFRROHXSMXFLSN-UHFFFAOYSA-N Glc6P Natural products OP(=O)(O)OCC(O)C(O)C(O)C(O)C=O VFRROHXSMXFLSN-UHFFFAOYSA-N 0.000 description 1
- 102000005548 Hexokinase Human genes 0.000 description 1
- 108700040460 Hexokinases Proteins 0.000 description 1
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 1
- 239000002202 Polyethylene glycol Substances 0.000 description 1
- 108700040099 Xylose isomerases Proteins 0.000 description 1
- PMMURAAUARKVCB-UHFFFAOYSA-N alpha-D-ara-dHexp Natural products OCC1OC(O)CC(O)C1O PMMURAAUARKVCB-UHFFFAOYSA-N 0.000 description 1
- WQZGKKKJIJFFOK-DVKNGEFBSA-N alpha-D-glucose Chemical compound OC[C@H]1O[C@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-DVKNGEFBSA-N 0.000 description 1
- 229940072107 ascorbate Drugs 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- KGBXLFKZBHKPEV-UHFFFAOYSA-N boric acid Chemical compound OB(O)O KGBXLFKZBHKPEV-UHFFFAOYSA-N 0.000 description 1
- 239000004327 boric acid Substances 0.000 description 1
- BVKZGUZCCUSVTD-UHFFFAOYSA-N carbonic acid Chemical compound OC(O)=O BVKZGUZCCUSVTD-UHFFFAOYSA-N 0.000 description 1
- 208000020832 chronic kidney disease Diseases 0.000 description 1
- 208000022831 chronic renal failure syndrome Diseases 0.000 description 1
- 238000004737 colorimetric analysis Methods 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 230000002526 effect on cardiovascular system Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 235000012209 glucono delta-lactone Nutrition 0.000 description 1
- 229960003681 gluconolactone Drugs 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 201000001421 hyperglycemia Diseases 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 229910000403 monosodium phosphate Inorganic materials 0.000 description 1
- 235000019799 monosodium phosphate Nutrition 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 230000001590 oxidative effect Effects 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 230000000291 postprandial effect Effects 0.000 description 1
- 125000004309 pyranyl group Chemical group O1C(C=CC=C1)* 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- AJPJDKMHJJGVTQ-UHFFFAOYSA-M sodium dihydrogen phosphate Chemical compound [Na+].OP(O)([O-])=O AJPJDKMHJJGVTQ-UHFFFAOYSA-M 0.000 description 1
- 238000013112 stability test Methods 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- UFTFJSFQGQCHQW-UHFFFAOYSA-N triformin Chemical compound O=COCC(OC=O)COC=O UFTFJSFQGQCHQW-UHFFFAOYSA-N 0.000 description 1
- 230000010245 tubular reabsorption Effects 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Images
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
- G01N21/31—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N2021/775—Indicator and selective membrane
Landscapes
- Physics & Mathematics (AREA)
- Chemical & Material Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Spectroscopy & Molecular Physics (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Plasma & Fusion (AREA)
- Engineering & Computer Science (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
本发明提供了一种检测1,5‑AG的试剂盒和检测方法。本发明第一方面提供了一种检测1,5‑AG的试剂盒,包括第一试剂和第二试剂,其中,所述第一试剂包括葡萄糖变构酶、葡萄糖氧化酶、过氧化物酶、抗坏血酸氧化酶、胆红素氧化酶、4‑AAP、表面活性剂、pH缓冲液、防腐剂以及稳定剂,所述第二试剂包括吡喃糖氧化酶、TOOS、pH缓冲液、防腐剂和稳定剂。采用本发明提供的试剂盒,能够利用紫外、可见光分析仪或者半自动/全自动生化分析仪检测出1,5‑AG的浓度,检测过程中不受葡萄糖,甘露糖,果糖等血清重糖类化合物的干扰,具有稳定性高、检测速度快、准确度高的优点。
Description
技术领域
本发明涉及临床生化诊断领域,尤其涉及一种检测1,5-AG的试剂盒和检测方法。
背景技术
1,5-脱水山梨醇(1,5-anhydroglucitol,1,5-AG),又叫做2-脱氧葡萄糖、1,5-脱水葡萄糖醇,分子式为C6H12O5(分子量164.2,CAS号为154-58-5),是一种具有吡喃环结构的六碳单糖,其结构与吡喃葡萄糖结构十分相似,主要区别在于吡喃葡萄糖C-1 位置上的羟基被氢所取代,形成C1椅形氧环,其结构决定了其具备较好的化学稳定性。
1,5-AG的浓度检测具有重要的临床意义,例如:正常情况下,血清中1,5-AG代谢稳定,浓度稳定在12-40mg/L,但当人体患糖尿病时,血清中1,5-AG的浓度会显著降低,主要原因是当人体患糖尿病时,大量葡萄糖由尿中排出竞争性的抑制了1,5-AG 的重吸收,从而使得1,5-AG大量经尿排出,导致血清中1,5-AG浓度降低,因此,血清中1,5-AG的浓度水平用于糖尿病诊断和疗效检测的常规指标;在非糖尿病人群中 1,5-AG浓度可以作为餐后血糖的筛查指标,监测餐后高血糖相关的心血管危险的发生;在慢性肾功能衰竭的病人的血清或尿液中1,5-AG的浓度也会明显下降,这是由于 1,5-AG重吸收减少造成的,血清或尿液中1,5-AG的浓度可很好的反映肾小管重吸收功能。
1,5-AG浓度的检测方法有很多,目前可实现自动分析的方法主要包括 ADP-HK-NADPH法和GK-PROD法,其中,ADP-HK-NADPH法原理主要为:1,5-AG 在ADP依赖性己糖激酶(ADP-HK)的作用下生成AG-6磷酸(AG-6-P),AG-6-P再和 NADP进行反应,生成NADPH,NADPH具有还原性,和显色剂作用生成显色物质,通过对显色物质进行比色,即可得知1,5-AG的浓度;GK-PROD法的原理主要为:使用吡喃糖氧化酶(PROD:EC1.1.3.10)、葡萄糖激酶(EC2.7.1.2)和三磷酸腺苷(ATP) 重生系统可检测1,5-AG的浓度变化,葡萄糖激酶和ATP重生系统可将葡萄糖转化为不与吡喃糖氧化酶反应的葡萄糖-6-磷酸,吡喃糖氧化酶氧化1,5-AG生成的过氧化氢可通过TRINDER反应进行检测,该方法可针对1,5-AG进行检测,可避免葡萄糖的干扰。
发明内容
有鉴于现有技术的上述缺陷,本发明所要解决的技术问题是提供一种检测1,5-AG的试剂盒和检测方法。
为实现上述目的,本发明第一方面提供了一种检测1,5-AG的试剂盒,包括第一试剂和第二试剂,其中,所述第一试剂包括葡萄糖变构酶、葡萄糖氧化酶、过氧化物酶、抗坏血酸氧化酶、胆红素氧化酶、4-AAP、表面活性剂、pH缓冲液、防腐剂以及稳定剂,所述第二试剂包括吡喃糖氧化酶、TOOS、pH缓冲液、防腐剂和稳定剂;
所述pH缓冲液控制所述第一试剂和第二试剂的pH为5.00-9.00。
进一步地,所述第一试剂中各组分的含量为:葡萄糖变构酶1-100KU/L、葡萄糖氧化酶1-100KU/L、过氧化物酶1-100KU/L、抗坏血酸氧化酶1-10008KU/L、胆红素氧化酶1-100KU/L、表面活性剂0.05-200g/L、pH缓冲液5-500mmol/L、稳定剂 0.05-200g/L、防腐剂0.05-10g/L;所述第二试剂中各组分的含量为:吡喃糖氧化酶 1-100KU/L、TOOS0.05-200g/L、表面活性剂0.05-200g/L、pH缓冲液5-500mmol/L、稳定剂0.05-200g/L、防腐剂0.05-10g/L。
进一步地,所述表面活性剂为曲拉通类、吐温类、布里杰类中的一种或多种。
进一步地,所述pH缓冲液为MES、MOPS、MOPSO、PIPES、HEPES、TES、 ADA、Tris、Bis-Tris、磷酸、柠檬酸、醋酸、碳酸、硼酸中的一种或多种,所述pH 缓冲液的浓度为5-500mmol/L。
进一步地,所述防腐剂为叠氮钠、PC300、山梨酸钾、新霉素、庆大霉素中的一种或多种。
进一步地,所述稳定剂为聚乙二醇类、多羟基醇类、金属离子络合剂中的一种或多种。
本发明第二方面提供了一种使用上述任一所述试剂盒的检测方法,包括如下检测步骤:
配制样品空白、样品、标准空白以及标准品,分别加入第一试剂,在30-37℃下反应,反应结束后,加入第二试剂,待反应结束后,测定相应的吸光度,计算得到样品中1,5-AG的浓度。
进一步地,根据式1计算所述样品中1,5-AG的浓度:
其中,A样品表示样品的吸光度、A样品空白表示样品空白的吸光度、A标准品表示标准品的吸光度、A标准空白表示标准空白的吸光度。
进一步地,测定吸光度时,波长为340-800nm。
进一步地,所述样品空白、样品、标准空白以及标准品的体积为1.5-50μL,所述第一试剂的体积为20-300μL,所述第二试剂的体积为20-300μL。
本发明提供了一种检测1,5-AG的试剂盒,其包括第一试剂和第二试剂,其检测原理如下:第一试剂中的葡萄糖变构酶将α-葡萄糖转化成β-D葡萄糖,β-D葡萄糖在葡萄糖氧化酶的作用下和氧气、水产生葡萄糖酸内酯和双氧水,双氧水被4-AAP和过氧化物酶转化成无色化合物,避免样品中内源性葡萄糖的干扰;随后加入第二试剂,吡喃糖氧化酶将1,5-AG氧化成1,5-脱水果糖和双氧水,随后双氧水、4-AAP、TOOS在过氧化物酶作用下产生红色醌亚胺,通过比色分析即可确定样本中1,5-AG的含量;此外,第一试剂中的抗坏血酸氧化酶用于消除样本中高抗坏血酸的影响,胆红素氧化酶用于消除样本中高胆红素的影响,表面活性剂用于消除样本中甘油三酯的影响。
本发明提供了一种检测1,5-AG的试剂盒和使用该试剂盒的检测方法,采用本发明提供的试剂盒,能够利用紫外、可见光分析仪或者半自动/全自动生化分析仪检测出 1,5-AG的浓度,检测过程中不受葡萄糖,甘露糖,果糖等血清重糖类化合物的干扰,具有稳定性高、检测速度快、准确度高的优点。
以下将结合具体实施例对本发明的构思、具体结构及产生的技术效果作进一步说明,以充分地了解本发明的目的、特征和效果。
附图说明
下面结合附图对本发明的具体实施方式作进一步详细的说明,其中:
图1为本发明配方配制试剂(研发试剂)和现有技术试剂(对照试剂)测定1,5AG 的结果相关性图。
具体实施方式
以下参考说明书附图介绍本发明的多个优选实施例,使其技术内容更加清楚和便于理解。本发明可以通过许多不同形式的实施例来得以体现,本发明的保护范围并非仅限于文中提到的实施例。
实施例1:
本实施例提供的试剂盒包括第一试剂和第二试剂,其中,
第一试剂(1L)包括:葡萄糖异构酶5KU、葡萄糖氧化酶10KU、抗坏血酸氧化酶10KU、胆红素氧化酶10KU、过氧化物酶5KU、4-AAP 5mmol、磷酸二氢钠 150mmol、曲拉通X100 20g、乙二胺四乙酸10mmol、叠氮钠5g;
第二试剂(1L)包括:吡喃糖氧化酶800KU、TOOS 5mmol、磷酸盐150mmol、曲拉通X100 20g、乙二胺四乙酸10mmol、叠氮钠5g。
(一)测试本发明实施例1提供的试剂盒的回收率,测试方法包括:
使用一个正常随机血清样本(记录为a),和一个高浓度的1,5-AG浓度样本 (记录为b),将样本a体积950μL和样本b体积50μL进行混合(记录为c);首先测试a样本浓度3次,随后测试样本b浓度3次,最后测试样本c浓度3次;三个样本浓度测试结果见表1:
表1三个样本浓度测试结果
a | b | c | |
1 | 75.62 | 218.56 | 82.56 |
2 | 75.73 | 218.15 | 82.71 |
3 | 75.45 | 218.34 | 82.18 |
均值 | 75.60 | 218.35 | 82.48 |
在血清样品中加入一定体积标准或校准品溶液(标准溶液体积与血清体积比应不大于1:20或其体积比不会产生基质变化,加入标准或校准品溶液后样品总浓度应在试剂盒测定线性范围内,标准品或校准品应有溯源性)或纯品,每个浓度重复测定3次,按式2计算回收率:
其中,R—回收率;
V-加入标准液体积;
V0-人血清样品的体积;
c-人血清样品加入标准液后的测定浓度;
c0-人血清样品的测定浓度;
ca-标准液的浓度。
根据式2计算得到回收率R为97.6%(一般行标回收率为90%-110%)。
(二)将实施例1提供的试剂盒与现有技术中北京九强提供的试剂盒进行比对:使用本发明配方配制试剂(研发试剂)和现有技术试剂(对照试剂)测定1,5 脱水山梨醇的结果相关性,相关性结果见图1,摘取50例血清样本数据见表2:
表2血清样本的测试数据
根据图1可知,相关性为0.999。
(三)将实施例1和北京九强提供的试剂在相同条件下进行37℃加速破坏试验, 10天加速稳定性数据见表3:
表3,研发试剂盒对照试剂的加速稳定性测试结果
天数 | 0 | 1 | 3 | 5 | 7 | 8 | 10 |
研发 | 100 | 99.6 | 101.2 | 98.7 | 98.2 | 97.5 | 96.8 |
对照 | 100 | 98.9 | 97.8 | 92.3 | 89.3 | 87.4 | 80.5 |
以上详细描述了本发明的较佳具体实施例。应当理解,本领域的普通技术无需创造性劳动就可以根据本发明的构思作出诸多修改和变化。因此,凡本技术领域中技术人员依本发明的构思在现有技术的基础上通过逻辑分析、推理或者有限的实验可以得到的技术方案,皆应在由权利要求书所确定的保护范围内。
Claims (10)
1.一种检测1,5-AG的试剂盒,其特征在于,包括第一试剂和第二试剂:
其中,所述第一试剂包括葡萄糖变构酶、葡萄糖氧化酶、过氧化物酶、抗坏血酸氧化酶、胆红素氧化酶、4-AAP、表面活性剂、pH缓冲液、防腐剂以及稳定剂;
所述第二试剂包括吡喃糖氧化酶、TOOS、pH缓冲液、防腐剂和稳定剂;
所述pH缓冲液控制所述第一试剂和第二试剂的pH为5.00-9.00。
2.如权利要求1所述的试剂盒,其特征在于,所述第一试剂中各组分的含量为:葡萄糖变构酶1-100KU/L、葡萄糖氧化酶1-100KU/L、过氧化物酶1-100KU/L、抗坏血酸氧化酶1-10008KU/L、胆红素氧化酶1-100KU/L、表面活性剂0.05-200g/L、pH缓冲液5-500mmol/L、稳定剂0.05-200g/L、防腐剂0.05-10g/L;所述第二试剂中各组分的含量为:吡喃糖氧化酶1-100KU/L、TOOS0.05-200g/L、表面活性剂0.05-200g/L、pH缓冲液5-500mmol/L、稳定剂0.05-200g/L、防腐剂0.05-10g/L。
3.如权利要求1或2所述的试剂盒,其特征在于,所述表面活性剂为曲拉通类、吐温类、布里杰类中的一种或多种。
4.如权利要求1或2所述的试剂盒,其特征在于,所述pH缓冲液为MES、MOPS、MOPSO、PIPES、HEPES、TES、ADA、Tris、Bis-Tris、磷酸、柠檬酸、醋酸、碳酸、硼酸中的一种或多种,所述pH缓冲液的浓度为5-500mmol/L。
5.如权利要求1或2所述的试剂盒,其特征在于,所述防腐剂为叠氮钠、PC300、山梨酸钾、新霉素、庆大霉素中的一种或多种。
6.如权利要求1或2所述的试剂盒,其特征在于,所述稳定剂为聚乙二醇类、多羟基醇类、金属离子络合剂中的一种或多种。
7.一种使用权利要求1-6任一项所述试剂盒的检测方法,其特征在于,包括如下检测步骤:
配制样品空白、样品、标准空白以及标准品,分别加入第一试剂,在30-37℃下反应,反应结束后,加入第二试剂,待反应结束后,测定相应的吸光度,计算得到样品中1,5-AG的浓度。
9.如权利要求7所述的检测方法,其特征在于,测定吸光度时,波长为340-800nm。
10.如权利要求7所述的检测方法,其特征在于,所述样品空白、样品、标准空白以及标准品的体积为1.5-50μL,所述第一试剂的体积为20-300μL,所述第二试剂的体积为20-300μL。
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202011363454.7A CN112630216A (zh) | 2020-11-27 | 2020-11-27 | 一种检测1,5-ag的试剂盒和检测方法 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202011363454.7A CN112630216A (zh) | 2020-11-27 | 2020-11-27 | 一种检测1,5-ag的试剂盒和检测方法 |
Publications (1)
Publication Number | Publication Date |
---|---|
CN112630216A true CN112630216A (zh) | 2021-04-09 |
Family
ID=75306514
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202011363454.7A Pending CN112630216A (zh) | 2020-11-27 | 2020-11-27 | 一种检测1,5-ag的试剂盒和检测方法 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN112630216A (zh) |
Citations (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US3964974A (en) * | 1972-09-28 | 1976-06-22 | Merck Patent Gesellschaft Mit Beschrankter Haftung | Enzymatic determination of glucose |
CN102154442A (zh) * | 2010-12-30 | 2011-08-17 | 北京九强生物技术股份有限公司 | 1,5-脱水山梨醇的检测方法和相关诊断试剂盒 |
CN102175670A (zh) * | 2010-12-30 | 2011-09-07 | 北京九强生物技术股份有限公司 | 血液1,5-脱水葡萄糖醇的检测方法及试剂盒 |
CN108949903A (zh) * | 2017-05-17 | 2018-12-07 | 广州市伊川生物科技有限公司 | 一种甘油三酯测定试剂盒及其测定方法 |
CN110702676A (zh) * | 2019-11-14 | 2020-01-17 | 北京华宇亿康生物工程技术有限公司 | 一种稳定性好的检测1,5-ag的试剂盒及方法 |
CN111808921A (zh) * | 2020-06-15 | 2020-10-23 | 武汉生之源生物科技股份有限公司 | 一种基于Trinder反应的检测试剂盒及其应用 |
-
2020
- 2020-11-27 CN CN202011363454.7A patent/CN112630216A/zh active Pending
Patent Citations (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US3964974A (en) * | 1972-09-28 | 1976-06-22 | Merck Patent Gesellschaft Mit Beschrankter Haftung | Enzymatic determination of glucose |
CN102154442A (zh) * | 2010-12-30 | 2011-08-17 | 北京九强生物技术股份有限公司 | 1,5-脱水山梨醇的检测方法和相关诊断试剂盒 |
CN102175670A (zh) * | 2010-12-30 | 2011-09-07 | 北京九强生物技术股份有限公司 | 血液1,5-脱水葡萄糖醇的检测方法及试剂盒 |
CN108949903A (zh) * | 2017-05-17 | 2018-12-07 | 广州市伊川生物科技有限公司 | 一种甘油三酯测定试剂盒及其测定方法 |
CN110702676A (zh) * | 2019-11-14 | 2020-01-17 | 北京华宇亿康生物工程技术有限公司 | 一种稳定性好的检测1,5-ag的试剂盒及方法 |
CN111808921A (zh) * | 2020-06-15 | 2020-10-23 | 武汉生之源生物科技股份有限公司 | 一种基于Trinder反应的检测试剂盒及其应用 |
Non-Patent Citations (5)
Title |
---|
侯振江编著: "《新编临床检验医学》", 31 March 2004, 军事医学科学出版社 * |
张宏萍: "重氮试剂消除胆红素对酶法测定甘油三酯的干扰", 《南通医学院学报》 * |
汪玄文: "胆红素氧化酶法消除胆红素对酶法测定胆固醇的干扰", 《医学检验与临床》 * |
王惠萱等: "血清尿酸自主研发生化诊断试剂的临床研究", 《国际检验医学杂志》 * |
郑铁生,鄢盛恺主编: "《临床生物化学检验》", 31 January 2020, 中国医药科技出版社 * |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Price et al. | A kinetic spectrophotometric assay for rapid determination of acetoacetate in blood. | |
EP2319937B1 (en) | Blood component measurement method utilizing hemolyzed whole blood, and kit for the method | |
CN102154442B (zh) | 1,5‑脱水山梨醇的检测方法和相关诊断试剂盒 | |
Fukumura et al. | Fully enzymatic method for determining 1, 5-anhydro-D-glucitol in serum | |
CN102435749B (zh) | 用循环酶法测定β-羟丁酸的液体稳定试剂盒 | |
CN102175670B (zh) | 血液1,5-脱水葡萄糖醇的检测方法及试剂盒 | |
Cooper | Methods for determining the amount of glucose in blood | |
CN105671127A (zh) | 一种稳定的酶法血清镁离子检测试剂盒 | |
CN107505273A (zh) | 血清总胆汁酸含量测定试剂盒及其使用方法 | |
CN109988816A (zh) | 一种腺苷脱氨酶测定试剂盒 | |
US5962248A (en) | Quantitative determination method for chloride ions | |
CN112255219A (zh) | 一种1,5-脱水山梨醇的测定试剂盒及其制备方法和应用 | |
CN111662955B (zh) | 一种腺苷脱氨酶测定试剂盒 | |
CN107653298A (zh) | 腺苷脱氨酶测定试剂盒 | |
CN112630216A (zh) | 一种检测1,5-ag的试剂盒和检测方法 | |
CN107254508B (zh) | 一种h2o2偶联的指示系统检测唾液酸的试剂盒 | |
EP1376133B1 (en) | Method of screening prediabetic state and screening reagent | |
CN111455020A (zh) | 一种1,5-脱水山梨醇检测试剂盒及检测方法 | |
CN109112181A (zh) | 一种5`-核苷酸酶活性测定方法及5`-核苷酸酶检测试剂盒 | |
CN112710656B (zh) | 一种测定葡萄糖含量的试剂盒及其应用 | |
Miskiewicz et al. | Evaluation of a glucose oxidase-peroxidase method adapted to the single-channel autoanalyzer and SMA 12/60 | |
WO2023206640A1 (zh) | 一种显色剂的稳定剂及稳定方法 | |
CN113655006B (zh) | 泌尿系结石成石危险因素检测试系统 | |
CN109541238A (zh) | 直接胆红素测定方法及试剂盒 | |
CN113984689A (zh) | 一种测定谷胱甘肽还原酶的试剂盒 |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination | ||
RJ01 | Rejection of invention patent application after publication | ||
RJ01 | Rejection of invention patent application after publication |
Application publication date: 20210409 |