CN100554948C - Homocysteine detection method and reagent thereof - Google Patents
Homocysteine detection method and reagent thereof Download PDFInfo
- Publication number
- CN100554948C CN100554948C CNB2004100167896A CN200410016789A CN100554948C CN 100554948 C CN100554948 C CN 100554948C CN B2004100167896 A CNB2004100167896 A CN B2004100167896A CN 200410016789 A CN200410016789 A CN 200410016789A CN 100554948 C CN100554948 C CN 100554948C
- Authority
- CN
- China
- Prior art keywords
- homocysteine
- adenosine
- reagent
- body fluid
- fluid sample
- 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.)
- Expired - Lifetime
Links
- FFFHZYDWPBMWHY-VKHMYHEASA-N L-homocysteine Chemical compound OC(=O)[C@@H](N)CCS FFFHZYDWPBMWHY-VKHMYHEASA-N 0.000 title claims abstract description 54
- 239000003153 chemical reaction reagent Substances 0.000 title claims abstract description 51
- 238000001514 detection method Methods 0.000 title description 4
- OIRDTQYFTABQOQ-KQYNXXCUSA-N adenosine Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O OIRDTQYFTABQOQ-KQYNXXCUSA-N 0.000 claims abstract description 44
- 238000000034 method Methods 0.000 claims abstract description 34
- 229960004452 methionine Drugs 0.000 claims abstract description 26
- 239000002126 C01EB10 - Adenosine Substances 0.000 claims abstract description 22
- 229960005305 adenosine Drugs 0.000 claims abstract description 22
- 210000001124 body fluid Anatomy 0.000 claims abstract description 18
- 239000010839 body fluid Substances 0.000 claims abstract description 18
- 230000000694 effects Effects 0.000 claims abstract description 15
- 238000003556 assay Methods 0.000 claims abstract description 10
- 230000004044 response Effects 0.000 claims abstract description 9
- 102000004190 Enzymes Human genes 0.000 claims description 24
- 108090000790 Enzymes Proteins 0.000 claims description 24
- 238000006243 chemical reaction Methods 0.000 claims description 21
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 claims description 16
- FFEARJCKVFRZRR-BYPYZUCNSA-N L-methionine Chemical compound CSCC[C@H](N)C(O)=O FFEARJCKVFRZRR-BYPYZUCNSA-N 0.000 claims description 16
- 229930182817 methionine Natural products 0.000 claims description 13
- ZKHQWZAMYRWXGA-KQYNXXCUSA-N Adenosine triphosphate 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-N 0.000 claims description 8
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 claims description 8
- 229910021529 ammonia Inorganic materials 0.000 claims description 8
- VHJLVAABSRFDPM-QWWZWVQMSA-N dithiothreitol Chemical compound SC[C@@H](O)[C@H](O)CS VHJLVAABSRFDPM-QWWZWVQMSA-N 0.000 claims description 8
- 101710169336 5'-deoxyadenosine deaminase Proteins 0.000 claims description 6
- 102000055025 Adenosine deaminases Human genes 0.000 claims description 6
- 101710088194 Dehydrogenase Proteins 0.000 claims description 5
- 101710101148 Probable 6-oxopurine nucleoside phosphorylase Proteins 0.000 claims description 4
- 102000030764 Purine-nucleoside phosphorylase Human genes 0.000 claims description 4
- 102100033220 Xanthine oxidase Human genes 0.000 claims description 4
- 108010093894 Xanthine oxidase Proteins 0.000 claims description 4
- 150000001875 compounds Chemical class 0.000 claims description 4
- BOPGDPNILDQYTO-NNYOXOHSSA-N nicotinamide-adenine dinucleotide Chemical compound C1=CCC(C(=O)N)=CN1[C@H]1[C@H](O)[C@H](O)[C@@H](COP(O)(=O)OP(O)(=O)OC[C@@H]2[C@H]([C@@H](O)[C@@H](O2)N2C3=NC=NC(N)=C3N=C2)O)O1 BOPGDPNILDQYTO-NNYOXOHSSA-N 0.000 claims description 4
- 239000000758 substrate Substances 0.000 claims description 4
- FFEARJCKVFRZRR-UHFFFAOYSA-N L-Methionine Natural products CSCCC(N)C(O)=O FFEARJCKVFRZRR-UHFFFAOYSA-N 0.000 claims description 3
- 229930195722 L-methionine Natural products 0.000 claims description 3
- 102000003992 Peroxidases Human genes 0.000 claims description 3
- 108040007629 peroxidase activity proteins Proteins 0.000 claims description 3
- JLVVSXFLKOJNIY-UHFFFAOYSA-N Magnesium ion Chemical compound [Mg+2] JLVVSXFLKOJNIY-UHFFFAOYSA-N 0.000 claims description 2
- 150000003838 adenosines Chemical class 0.000 claims description 2
- 229910001425 magnesium ion Inorganic materials 0.000 claims description 2
- 238000006911 enzymatic reaction Methods 0.000 claims 1
- 230000003301 hydrolyzing effect Effects 0.000 claims 1
- 102000005234 Adenosylhomocysteinase Human genes 0.000 abstract description 14
- 108020002202 Adenosylhomocysteinase Proteins 0.000 abstract description 14
- 238000005516 engineering process Methods 0.000 abstract description 5
- 230000035945 sensitivity Effects 0.000 abstract description 5
- 230000002255 enzymatic effect Effects 0.000 abstract description 3
- 238000005259 measurement Methods 0.000 abstract description 2
- 230000002829 reductive effect Effects 0.000 description 9
- 239000005515 coenzyme Substances 0.000 description 7
- RLFWWDJHLFCNIJ-UHFFFAOYSA-N 4-aminoantipyrine Chemical compound CN1C(C)=C(N)C(=O)N1C1=CC=CC=C1 RLFWWDJHLFCNIJ-UHFFFAOYSA-N 0.000 description 6
- 238000007254 oxidation reaction Methods 0.000 description 6
- 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 5
- 210000004369 blood Anatomy 0.000 description 5
- 239000008280 blood Substances 0.000 description 5
- 239000003638 chemical reducing agent Substances 0.000 description 5
- 230000004087 circulation Effects 0.000 description 5
- FFFHZYDWPBMWHY-UHFFFAOYSA-N L-Homocysteine Natural products OC(=O)C(N)CCS FFFHZYDWPBMWHY-UHFFFAOYSA-N 0.000 description 4
- 108010049351 adenosine nucleosidase Proteins 0.000 description 4
- 239000003795 chemical substances by application Substances 0.000 description 4
- 239000003593 chromogenic compound Substances 0.000 description 4
- FDGQSTZJBFJUBT-UHFFFAOYSA-N hypoxanthine Chemical compound O=C1NC=NC2=C1NC=N2 FDGQSTZJBFJUBT-UHFFFAOYSA-N 0.000 description 4
- 229930027945 nicotinamide-adenine dinucleotide Natural products 0.000 description 4
- 230000003647 oxidation Effects 0.000 description 4
- 239000008363 phosphate buffer Substances 0.000 description 4
- 238000013016 damping Methods 0.000 description 3
- 239000012530 fluid Substances 0.000 description 3
- 230000009467 reduction Effects 0.000 description 3
- 238000007616 round robin method Methods 0.000 description 3
- -1 sulfhydryl compound Chemical class 0.000 description 3
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 2
- 102100036664 Adenosine deaminase Human genes 0.000 description 2
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 2
- 239000007995 HEPES buffer Substances 0.000 description 2
- UGQMRVRMYYASKQ-UHFFFAOYSA-N Hypoxanthine nucleoside Natural products OC1C(O)C(CO)OC1N1C(NC=NC2=O)=C2N=C1 UGQMRVRMYYASKQ-UHFFFAOYSA-N 0.000 description 2
- XUJNEKJLAYXESH-REOHCLBHSA-N L-Cysteine Chemical compound SC[C@H](N)C(O)=O XUJNEKJLAYXESH-REOHCLBHSA-N 0.000 description 2
- WHUUTDBJXJRKMK-VKHMYHEASA-N L-glutamic acid Chemical compound OC(=O)[C@@H](N)CCC(O)=O WHUUTDBJXJRKMK-VKHMYHEASA-N 0.000 description 2
- 229930195725 Mannitol Natural products 0.000 description 2
- 102000007357 Methionine adenosyltransferase Human genes 0.000 description 2
- 108010007784 Methionine adenosyltransferase Proteins 0.000 description 2
- LCTONWCANYUPML-UHFFFAOYSA-N Pyruvic acid Chemical compound CC(=O)C(O)=O LCTONWCANYUPML-UHFFFAOYSA-N 0.000 description 2
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 2
- OUUQCZGPVNCOIJ-UHFFFAOYSA-M Superoxide Chemical class [O-][O] OUUQCZGPVNCOIJ-UHFFFAOYSA-M 0.000 description 2
- 239000013504 Triton X-100 Substances 0.000 description 2
- 229920004890 Triton X-100 Polymers 0.000 description 2
- PTKRUDMLGIIORX-ITGWJZMWSA-N [[(2r,3r,4r,5r)-5-(6-aminopurin-9-yl)-3-hydroxy-4-phosphonooxyoxolan-2-yl]methoxy-hydroxyphosphoryl] [(2r,3s,4r,5r)-5-(3-carbamoyl-4h-pyridin-1-yl)-3,4-dihydroxyoxolan-2-yl]methyl hydrogen phosphate;cyclohexanamine Chemical compound NC1CCCCC1.NC1CCCCC1.NC1CCCCC1.NC1CCCCC1.C1=CCC(C(=O)N)=CN1[C@H]1[C@H](O)[C@H](O)[C@@H](COP(O)(=O)OP(O)(=O)OC[C@@H]2[C@H]([C@@H](OP(O)(O)=O)[C@@H](O2)N2C3=NC=NC(N)=C3N=C2)O)O1 PTKRUDMLGIIORX-ITGWJZMWSA-N 0.000 description 2
- 238000002835 absorbance Methods 0.000 description 2
- 230000003197 catalytic effect Effects 0.000 description 2
- 210000004027 cell Anatomy 0.000 description 2
- 235000010355 mannitol Nutrition 0.000 description 2
- 239000000594 mannitol Substances 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 230000004060 metabolic process Effects 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 235000018102 proteins Nutrition 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- MWFOPMKUGZLPQA-UHFFFAOYSA-M sodium;3-(n-ethyl-3-methoxyanilino)propane-1-sulfonate Chemical compound [Na+].[O-]S(=O)(=O)CCCN(CC)C1=CC=CC(OC)=C1 MWFOPMKUGZLPQA-UHFFFAOYSA-M 0.000 description 2
- 229910052717 sulfur Inorganic materials 0.000 description 2
- 239000011593 sulfur Substances 0.000 description 2
- RTDXMJJSUVCSKB-RUCXOUQFSA-N (2s)-2-azanyl-4-sulfanyl-butanoic acid Chemical compound OC(=O)[C@@H](N)CCS.OC(=O)[C@@H](N)CCS RTDXMJJSUVCSKB-RUCXOUQFSA-N 0.000 description 1
- IHPYMWDTONKSCO-UHFFFAOYSA-N 2,2'-piperazine-1,4-diylbisethanesulfonic acid Chemical compound OS(=O)(=O)CCN1CCN(CCS(O)(=O)=O)CC1 IHPYMWDTONKSCO-UHFFFAOYSA-N 0.000 description 1
- YDBHVMTTYXWHLI-UHFFFAOYSA-N 2,4,6-tribromo-3-hydroxybenzoic acid Chemical compound OC(=O)C1=C(Br)C=C(Br)C(O)=C1Br YDBHVMTTYXWHLI-UHFFFAOYSA-N 0.000 description 1
- MSUBEYXGDSJIPS-UHFFFAOYSA-N 3-(2-methylanilino)propan-1-ol Chemical compound CC1=CC=CC=C1NCCCO MSUBEYXGDSJIPS-UHFFFAOYSA-N 0.000 description 1
- BTIDJAQNJLWPTI-UHFFFAOYSA-N 3-(n-ethyl-3,5-dimethoxyanilino)-2-hydroxypropane-1-sulfonic acid Chemical compound OS(=O)(=O)CC(O)CN(CC)C1=CC(OC)=CC(OC)=C1 BTIDJAQNJLWPTI-UHFFFAOYSA-N 0.000 description 1
- 102000015081 Blood Coagulation Factors Human genes 0.000 description 1
- 108010039209 Blood Coagulation Factors Proteins 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- GKGQYWDLSLRTPJ-UHFFFAOYSA-N C(=O)O.OC=1C(=CC(=CC1Br)Br)Br Chemical compound C(=O)O.OC=1C(=CC(=CC1Br)Br)Br GKGQYWDLSLRTPJ-UHFFFAOYSA-N 0.000 description 1
- XATCVRKQPMGQFW-UHFFFAOYSA-N CCN(CC(CS(=O)(=O)O)O)C1=CC=CC=C1OC Chemical compound CCN(CC(CS(=O)(=O)O)O)C1=CC=CC=C1OC XATCVRKQPMGQFW-UHFFFAOYSA-N 0.000 description 1
- VEIZTCQKWBVPBD-UHFFFAOYSA-N CCN(CCCS(=O)(=O)O)C1=CC=CC=C1OC Chemical compound CCN(CCCS(=O)(=O)O)C1=CC=CC=C1OC VEIZTCQKWBVPBD-UHFFFAOYSA-N 0.000 description 1
- 208000024172 Cardiovascular disease Diseases 0.000 description 1
- 108010001336 Horseradish Peroxidase Proteins 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 1
- 206010020608 Hypercoagulation Diseases 0.000 description 1
- UGQMRVRMYYASKQ-KQYNXXCUSA-N Inosine Chemical compound O[C@@H]1[C@H](O)[C@@H](CO)O[C@H]1N1C2=NC=NC(O)=C2N=C1 UGQMRVRMYYASKQ-KQYNXXCUSA-N 0.000 description 1
- 229930010555 Inosine Natural products 0.000 description 1
- XYFCBTPGUUZFHI-UHFFFAOYSA-N Phosphine Chemical compound P XYFCBTPGUUZFHI-UHFFFAOYSA-N 0.000 description 1
- 102000004160 Phosphoric Monoester Hydrolases Human genes 0.000 description 1
- 108090000608 Phosphoric Monoester Hydrolases Proteins 0.000 description 1
- 208000007536 Thrombosis Diseases 0.000 description 1
- LEHOTFFKMJEONL-UHFFFAOYSA-N Uric Acid Chemical compound N1C(=O)NC(=O)C2=C1NC(=O)N2 LEHOTFFKMJEONL-UHFFFAOYSA-N 0.000 description 1
- TVWHNULVHGKJHS-UHFFFAOYSA-N Uric acid Natural products N1C(=O)NC(=O)C2NC(=O)NC21 TVWHNULVHGKJHS-UHFFFAOYSA-N 0.000 description 1
- 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 1
- 239000012190 activator Substances 0.000 description 1
- 125000002015 acyclic group Chemical group 0.000 description 1
- 229940024606 amino acid Drugs 0.000 description 1
- 235000001014 amino acid Nutrition 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000002421 anti-septic effect Effects 0.000 description 1
- 230000003143 atherosclerotic effect Effects 0.000 description 1
- 208000022379 autosomal dominant Opitz G/BBB syndrome Diseases 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 230000017531 blood circulation Effects 0.000 description 1
- 239000003114 blood coagulation factor Substances 0.000 description 1
- 229940098773 bovine serum albumin Drugs 0.000 description 1
- 230000005779 cell damage Effects 0.000 description 1
- 208000037887 cell injury Diseases 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000004737 colorimetric analysis Methods 0.000 description 1
- 239000008139 complexing agent Substances 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 230000008878 coupling Effects 0.000 description 1
- 238000010168 coupling process Methods 0.000 description 1
- 238000005859 coupling reaction Methods 0.000 description 1
- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 description 1
- 235000018417 cysteine Nutrition 0.000 description 1
- 238000007357 dehydrogenase reaction Methods 0.000 description 1
- 230000017858 demethylation Effects 0.000 description 1
- 238000010520 demethylation reaction Methods 0.000 description 1
- 238000012631 diagnostic technique Methods 0.000 description 1
- 238000002875 fluorescence polarization Methods 0.000 description 1
- 229960002989 glutamic acid Drugs 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 238000003018 immunoassay Methods 0.000 description 1
- 229960003786 inosine Drugs 0.000 description 1
- 230000002452 interceptive effect Effects 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 150000001455 metallic ions Chemical class 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 239000000049 pigment Substances 0.000 description 1
- 229940107700 pyruvic acid Drugs 0.000 description 1
- HDARHUHTZKLJET-UHFFFAOYSA-M sodium;3-(n-ethyl-3,5-dimethoxyanilino)-2-hydroxypropane-1-sulfonate Chemical compound [Na+].[O-]S(=O)(=O)CC(O)CN(CC)C1=CC(OC)=CC(OC)=C1 HDARHUHTZKLJET-UHFFFAOYSA-M 0.000 description 1
- CJUDSKIRZCSXJA-UHFFFAOYSA-M sodium;3-(n-ethyl-3-methoxyanilino)-2-hydroxypropane-1-sulfonate Chemical compound [Na+].[O-]S(=O)(=O)CC(O)CN(CC)C1=CC=CC(OC)=C1 CJUDSKIRZCSXJA-UHFFFAOYSA-M 0.000 description 1
- 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 description 1
- 239000004094 surface-active agent Substances 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 201000005665 thrombophilia Diseases 0.000 description 1
- 229940116269 uric acid Drugs 0.000 description 1
- 210000003556 vascular endothelial cell Anatomy 0.000 description 1
Images
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention relates to a kind of enzymatic determination method and reagent thereof that adopts circular increment technology, be used for measuring the content of body fluid sample homocysteine.Be characterized in:, improve and measure sensitivity, thereby can carry out automatic assay to the homocysteine in the body fluid sample as common zymetology diagnostic reagent by adopting a kind of circular increment technology.Homocysteine in the body fluid sample under the effect of HcyMetase and S-Adenosylhomocysteine synthase with S-adenosine-L-methionine circular response repeatedly, produce adenosine, the speed that produces adenosine is directly proportional with the content of homocysteine in the sample, by measuring the generating rate of adenosine, just can reach the purpose of measuring homocysteine content in the body fluid sample.Adopt the quantitative measurement reagent of this method preparation to have convenient, fast, robotization and highly sensitive characteristics.
Description
Technical field:
The present invention relates to the assay method and the reagent thereof of homocysteine
Background technology:
Homocysteine (Homocysteine) is a sulfur-containing amino acid, is generated by the methionine demethylation in cell.Recent studies show that, homocysteine is by producing superoxides and superoxide vascular endothelial cell injury, change the clotting factor function, increase thrombophilia, promote atherosclerotic and thrombosis, cardiovascular disease incidence rate and mortality ratio are increased, and meaning is very important clinically therefore to measure the concentration of homocysteine in the blood.When homocysteine in the cell inner accumulated, and after entering blood circulation, most ofly exist with oxidized form, and and protein combination, reduction only accounts for 1% just as the type halfcystine in blood.Measure the content of total homocysteine in the blood, generally need to adopt reductive agent that the oxidized form homocysteine is reduced to the reduced form homocysteine.
The assay method of homocysteine in clinical practice mainly comprises: high performance liquid chromatography, enzyme immunoassay (EIA), fluorescence polarization method etc.The common feature of said method is to need special determining instrument, complicated operation, tests consuming time longlyer, can not be applied to the automated analysis of clinical big flow, and cost an arm and a leg.
Because the content of homocysteine in sample is lower, the assay method of routine clinical zymetology can't reach required mensuration sensitivity, has limited the employing of conventional method.Measure the technical elements of homocysteine at Enzymology method, (referring to United States Patent (USP) #6686172) such as Naoto Matsuyama invented a kind of acyclic enzymatic determination technology of utilizing HcyMetase and S-Adenosylhomocysteine synthase, improve and measure sensitivity of method, but this method complex operation, the reagent complexity, need before measuring to add sulfhydryl compound, must carry out multinomial blank determination simultaneously during mensuration, increased the inaccuracy of assay method.
The invention discloses the enzyme round-robin method of homocysteine in a kind of new mensuration body fluid sample, this method is not subjected to the interference of interfering material in the body fluid sample, has good reaction sensitivity.And adopt first and measure toolenzyme commonly used in the homocysteine method, as: HcyMetase and SAHH are used for the enzyme circular increment and measure.In addition, the present invention also discloses based on the enzyme round-robin method of above-mentioned two kinds of toolenzymes simultaneously first and has utilized the homocysteine detection reagent of this method preparation, this reagent can be applied to widely used clinical automatic analyzer at present, thereby reaches the requirement of extensive mensuration sample.
Summary of the invention:
The invention provides a kind of enzymatic determination method and reagent thereof that adopts circular increment technology, measure the content of homocysteine in the body fluid sample.This circular increment is measured system and mainly is made up of HcyMetase and S-Adenosylhomocysteine synthase.Oxidized form homocysteine in the body fluid sample under the catalytic action of HcyMetase (EC2.1.1.10), with S-adenosine-L-methionine reaction, generates S-adenosine-L-homocysteine and L-methionine after the reductive agent reduction.S-adenosine-L-homocysteine is hydrolyzed to L-homocysteine and adenosine (seeing figure) under the effect of S-Adenosylhomocysteine synthase (EC 3.3.1.1).Therefore, L-homocysteine in the sample is circular response repeatedly, constantly produces adenosine, and the generating rate of adenosine is directly proportional with the content of homocysteine in the sample, by measuring the generating rate of adenosine, just can reach the purpose of measuring homocysteine content in the body fluid sample.
The assay method of adenosine is a lot, only enumerated the auxiliary enzymes system of two kinds of common mensuration adenosines among the present invention: (1) adenosine generates ammonia and inosine under the effect of adenosine deaminase, the latter is through the purine nucleoside phosphorylase effect, generate hypoxanthine with phosphatase reaction, under the effect of xanthine oxidase, hypoxanthine finally is oxidized to uric acid, and produces superoxide hydrogen, through reacting, measure the pigment that generates with chromogenic compound; Perhaps (2) adopt the method for various mensuration ammonia, as the glutamte dehydrogenase reaction method.Ammonia and α-Tong Wuersuan produce L-glutamic acid under the effect of glutamte dehydrogenase, coenzyme NAD H or NADPH are oxidized to NAD simultaneously
+Or NADP
+, reactive system descends at the absorbance at 340nm place.Participate in reaction because the homocysteine in the sample circulates repeatedly, constantly produce adenosine, so improved the sensitivity of measuring greatly.
Description of drawings:
The methodology principle of invention: the oxidized form homocysteine (Hcy) in the sample is after the reducing agent reduction, under the catalytic action of HcyMetase (HcyMetase), with SAM (AdoMet) reaction, generate S-adenosine-L-homocysteine (AdoHcy) and L-Methionine (Met). S-adenosine-L-homocysteine is hydrolyzed to L-homocysteine and adenosine (Ado) under the effect of S-Adenosylhomocysteine synthase (AdoHcyase). Generate SAM (AdoMet) under the methionine that reaction generates and the atriphos in the reagent (ATP) turn adenosinase (MAT) at methionine the effect. L-homocysteine iterative cycles reaction in the sample, constantly produce adenosine, the generating rate of adenosine is directly proportional with the content of homocysteine in the sample, by measuring the generating rate of adenosine, just can reach the purpose of measuring content of homocysteine in the body fluid sample.
Most of homocysteine all exists with oxidized form in blood, wherein 80-90% and protein combination, 5-10% and homocysteine self combination, the in addition combination such as 5-10% and cysteine formation mixed type homocysteine disulphide; Also original shape only accounts for about 1%. When measuring in the blood total homocysteine content, generally adopt reducing agent to reduce the disulfide bond of its combination, it is existed with free reduced form homocysteine form. Chemical reducing agent commonly used has dithiothreitol (DTT) (DTT), three-(carboxyethyl) hydrogen phosphide hydrochloride (TCEP) etc. The consumption of reducing agent is unsuitable excessive, in order to avoid the interference measurement reaction, such as the consumption of DTT preferably less than 10mM.
In addition, another effect of sulfur-bearing reducing agent is the accelerative activator that turns adenosinase as HcyMetase and methionine.
S-adenosine-L-methionine is as the initial compounds of enzyme circular response, can adopt finished commercial prod, but because common commercial grade instability, adopt the auxiliary enzymes system to generate among the present invention, promptly under the effect of methionine adenosyltransferase, generate S-adenosine-L-methionine by methionine and ATP, another advantage of introducing this auxiliary enzymes system is further to promote desirable the Direction of Reaction in the major cycle reaction, and can reduce the reagent dosage of methionine.This auxiliary enzymes system is not essential in assay method is learned, and measures as long as can provide stable S-adenosine-L-methionine to be used for circulation, and any enzyme system or other method all are feasible.
The present invention has created the enzyme circular response of being made up of HcyMetase and S-Adenosylhomocysteine synthase first, and is applied to the mensuration of homocysteine.Based on the characteristics that the enzyme process circular increment is measured, the enzyme use amount of circulating reaction system does not require very high, and end reaction concentration generally between 0.1～50ku/L, is preferably between 0.2～10ku/L.Certainly, the consumption that increases enzyme does not influence the application of the inventive method, but can increase the manufacturing cost of reagent.Optimize enzyme and substrate application quantity and can promote circular response to carry out to desirable direction, ATP should be excessive in circulative metabolism, and greater than the consumption of methionine, usable range generally between 0.1～90mM/L, is preferably within 1～80mM/L scope.The consumption of S-Adenosylhomocysteine synthase should be greater than the consumption of HcyMetase, and the consumption of methionine adenosyltransferase is more preferably greater than the consumption of HcyMetase, and in addition, adenosine deaminase should be excessive.
The present invention discloses a kind of assay method, this method is not subjected to the interference of materials such as adenosine in the body fluid sample, methionine, cystathionie.
The present invention also provides two kinds of enzyme circular responses of being made up of HcyMetase and S-Adenosylhomocysteine synthase to measure reagent simultaneously first, is used for measuring the content of body fluid sample homocysteine.
Utilize enzyme round-robin method of the present invention, can prepare the diagnostic reagent of various homocysteines according to common zymetology diagnostic techniques.In one embodiment of the invention, adopt peroxidase colorimetric method (being also referred to as Trinder ' s method), the adenosine that the circulation system produces is under the further effect of adenosine deaminase, purine nucleoside phosphorylase, xanthine oxidase, generate hydrogen peroxide, under the effect of peroxidase, hydrogen peroxide and 4-amino-antipyrine and the reaction of oxidation chromogenic compound, chromogenesis.By detecting pigmentogenic speed, can calculate the content of homocysteine in the sample.Measure in the reagent at clinical enzymology, oxidation chromogenic compound commonly used is a lot, as: N-ethyl-N-sulfo group hydroxypropyl--toluidine (TOOS), 3-hydroxyl-2,4,6-tribromo-benzene formic acid (TBHBA), N-ethyl-N-(3-sulfopropyl)--methyl oxyaniline (ADPS), N-ethyl-N-(2-hydroxyl-3-sulfopropyl)--methyl oxyaniline (ADOS), N-ethyl-N-(2-hydroxyl-3-sulfopropyl)-3,5-dimethoxy aniline (DAOS) etc.Also have another kind of oxidation color-producing bodies not need the coupling of 4-amino-antipyrine.The consumption of oxidation chromogenic compound is preferably in 0.5～3mM/L generally at 0.01～10mM/L.
In another embodiment of the present invention, adopt a kind of Enzymology method of measuring ammonia.The adenosine that the circulation system produces produces ammonia under the effect of adenosine deaminase, under the further effect of glutamte dehydrogenase auxiliary measuring system, reduced coenzyme (as: NADH, NADPH, thio-NADH, thio-NADPH etc.) is oxidized to oxidized coenzyme, by the speed that monitoring 340nm place absorbance value descends, can calculate the content of homocysteine in the sample.The consumption of reduced coenzyme is preferably in 0.15～0.4mM/L generally at 0.1～0.8mM/L.
Should comprise damping fluid in the homocysteine detection reagent that adopts circulative metabolism of the present invention to prepare, as: phosphate buffer, N-(2-hydroxyethyl) piperazine-N '-(2-ethylsulfonic acid) be damping fluid etc. (HEPES), metallic ion, as the essential composition of reactions such as magnesium ion, also can contain compounds such as surfactant, complexing agent (as: EDTA) and antiseptic.
The assay method of above-mentioned two kinds of reagent, identical with application process conventional in this specialty, as: can adopt rate method or end-point method,, calculate the content of measured object, not repeat them here by reference calibration object or drawing standard concentration curve etc.Those skilled in the art can make various similar mensuration reagent according to principle of the present invention and method, but does not detach principle of the present invention and range of application.
Specific embodiment:
Embodiment 1: adopt the enzyme circular response of forming by HcyMetase and S-Adenosylhomocysteine synthase, the hydrogen peroxide color producing reaction that the mensuration reagent system produces (Trinder ' the s reaction).
Reagent 1:(R1: R2=3: 1)
| Reagent components | Every liter of consumption | Every liter of amount ranges commonly used |
| Phosphate buffer, pH7.0,37 ℃ | 100mM | 10～300mM |
| EDTA.2Na | 0.2mM | 0.1～20mM |
| MgSO 4 | 15mM | 0.5～100mM |
| Triton X-100 | 0.1％ | 0.01～5％ |
| DTT | 2mM | 0.1～20mM |
| ATP | 80mM | 0.1～90mM |
| ADA | 3ku | 0.1～200ku |
| ADPS | 2mM | 0.01～20mM |
| AdoHcyase | 10ku | 0.1～50ku |
| Mannitol | 20mM | 1～100mM |
| Oxidized coenzyme NAD | 0.1mM | 0.01～20mM |
| Methionine changes adenosinase | 15ku | 0.1～50ku |
| Purine nucleoside phosphorylase | 2ku | 0.1～50ku |
| Xanthine oxidase | 3ku | 0.1～50ku |
| Horseradish peroxidase | 2ku | 0.1～50ku |
| HcyMetase | 4ku | 0.1～100ku |
Reagent 2:(R1: R2=3: 1)
| Reagent components | Every liter of consumption | Every liter of amount ranges commonly used |
| Phosphate buffer, pH7.0,37 ℃ | 100mM | 10～300mM |
| EDTA.2Na | 0.2mM | 0.1～20mM |
| MgSO 4 | 15mM | 0.5～100mM |
| Triton X-100 | 0.1％ | 0.01～5％ |
| Methionine | 0.5mM | 0.1～90mM |
| The 4-amino-antipyrine | 1mM |
Embodiment 1 reagent 1 prescription is used for reduction-oxidation type homocysteine, gets rid of the interference of adenosine in the sample, and reagent 1 complete enzyme circulation is simultaneously measured reagent and is used to get rid of subsidiary reaction.When measuring sample, adopt fixed time, the ratio of reagent and sample is fixedly requirement not, but same mensuration batch planted agent's unanimity.As reagent 1: sample: reagent 2 is 300: 20: 100,37 ℃ of temperature, measure wavelength 540nm, reagent 1 was hatched 300 seconds in the mensuration temperature after adding sample or calibration object, and the adenosine of removing in the sample disturbs, and added reagent 2 then and began to measure, 0～120 second time delay, can be used for getting rid of the subsidiary reaction of other S-adenosine-L-methionine → S-adenosine-L-homocysteine in the sample, minute 180～300 seconds, reading are selected at least 2 available points in minute.
Embodiment 2: adopt the enzyme circular response of being made up of HcyMetase and S-Adenosylhomocysteine synthase, by the ammonia reagent method, measure the oxidized speed of reduced coenzyme in the set time section.
Reagent 1:(R1: R2=4: 1)
| Reagent components | Every liter of consumption | Every liter of amount ranges commonly used |
| Phosphate buffer, pH6.5,37 ℃ | 150mM | 500～500mM |
| EDTA.2Na | 0.5mM | 0.1～20mM |
| MgSO 4 | 15mM | 0.5～100mM |
| DTT | 1.5mM | 0.1～20mM |
| ATP | 80mM | 0.1～90mM |
| ADA | 3ku | 0.1～200ku |
| AdoHcyase | 10ku | 0.1～50ku |
| Bovine serum albumin(BSA) | 0.2％ | 0.01～10％ |
| α-Tong Wuersuan | 7.5mM | 1～20mM |
| Methionine changes adenosinase | 15ku | 0.1～100ku |
| Glutamte dehydrogenase | 5ku | 1～50ku |
| HcyMetase | 5ku | 0.1～50ku |
Reagent 2:(R1: R2=4: 1)
| Reagent components | Every liter of consumption | Every liter of amount ranges commonly used |
| The HEPES damping fluid, pH8.3,37 ℃ | 50mM | 10～200mM |
| EDTA.2Na | 0.5mM | 0.1～20mM |
| MgSO 4 | 15mM | 0.5～100mM |
| Methionine | 10mM | 0.1～90mM |
| Mannitol | 20mM | 1～100mM |
| Reduced coenzyme NADH | 0.8mM | 0.1～10mM |
| Lactic dehydrogenase | 2ku | 0.5～20ku |
Embodiment 2 reagent 1 prescription is used for reduction-oxidation type homocysteine, and reagent 2 is combined into complete enzyme circulation with reagent 1 and measures reagent.When measuring sample, adopt fixed time, the ratio of reagent and sample is fixedly requirement not, but same mensuration batch planted agent's unanimity.As reagent 1: sample: reagent 2 is 200: 25: 50,37 ℃ of temperature, measure wavelength 340nm, reagent 1 was hatched 300 seconds in the mensuration temperature after adding sample or calibration object, added reagent 2 then, postponed 60～120 seconds, the interference of ammonia and pyruvic acid in the eliminating sample, and the subsidiary reaction of other S-adenosine-L-methionine → S-adenosine-L-homocysteine, minute 180 seconds, reading are selected at least 2 available points in minute.
The compound method of the foregoing description reagent only is used to illustrate principle of the present invention and application thereof, and the present invention never is confined to above-mentioned range of application of giving an example; In addition, the professional and technical personnel in association area of the present invention can make similar with it various mensuration reagent according to principle of the present invention and method, but not detach principle of the present invention and range of application.
Claims (6)
1, a kind of method of measuring homocysteine in the body fluid sample, it is characterized in that: (a) adopt by S-adenosine-L-methionine: the enzyme circular response that L-halfcystine S-transmethylase and S-adenosine-L-homocysteine hydrolytic enzyme and substrate thereof are formed, the homocysteine that contains in the tested sample produces adenosine under the effect of this enzyme circulating reaction system; (b) speed of this adenosine generation is directly proportional with the content of homocysteine in the sample, by measuring the generating rate of adenosine, can obtain the content of homocysteine in the sample.
2, the method for homocysteine in the mensuration body fluid sample according to claim 1, wherein substrate is meant S-adenosine-L-methionine.
3, the method for homocysteine in the mensuration body fluid sample according to claim 1, its employed reagent also comprises: dithiothreitol (DTT) and magnesium ion.
4, the method for homocysteine in the mensuration body fluid sample according to claim 2, be to adopt hydrogen peroxide color producing reaction method, its reagent also should comprise: adenosine deaminase, purine nucleoside phosphorylase, xanthine oxidase, peroxidase and Trinde ' s chromogen compound.
5, the method for homocysteine in the mensuration body fluid sample according to claim 2 is to adopt the ammonia assay method, and its reagent also comprises: adenosine deaminase, glutamte dehydrogenase, α-Tong Wuersuan and reduced diphosphopyridine nucleotide or reduced diphosphopyridine nucleotide I.
6, the method for homocysteine in the mensuration body fluid sample according to claim 2, wherein substrate S-adenosine-L-methionine is by atriphos, methionine and atriphos: the enzymatic reaction system that L-methionine S-adenylase is formed is generated.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB2004100167896A CN100554948C (en) | 2004-03-08 | 2004-03-08 | Homocysteine detection method and reagent thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB2004100167896A CN100554948C (en) | 2004-03-08 | 2004-03-08 | Homocysteine detection method and reagent thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1560610A CN1560610A (en) | 2005-01-05 |
| CN100554948C true CN100554948C (en) | 2009-10-28 |
Family
ID=34440654
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNB2004100167896A Expired - Lifetime CN100554948C (en) | 2004-03-08 | 2004-03-08 | Homocysteine detection method and reagent thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN100554948C (en) |
Families Citing this family (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US7097968B2 (en) | 2003-07-10 | 2006-08-29 | General Atomics | Methods and compositions for assaying homocysteine |
| US8476034B2 (en) | 2003-07-10 | 2013-07-02 | General Atomics | Methods and compositions for assaying homocysteine |
| CN101915760B (en) * | 2010-07-07 | 2013-10-09 | 东华大学 | Method for detecting cysteine content in real time by colorimetric method |
| CN102243227B (en) * | 2010-12-16 | 2014-01-15 | 浙江亚太药业股份有限公司 | Measuring method of asymmetric dimethylarginine concentration and measuring reagent thereof |
| CN102766677A (en) * | 2012-07-31 | 2012-11-07 | 武汉生之源生物科技有限公司 | Lactic dehydrogenase detection kit and preparation method thereof |
| CN103913581A (en) * | 2013-11-20 | 2014-07-09 | 天津市宝坻区人民医院 | Cyclophorase determination method for triglyceride in serum |
| CN103954675B (en) * | 2014-05-06 | 2016-01-20 | 济南大学 | A kind of preparation method of S-adenosylmethionine molecular engram sensor and application |
| CN104111338B (en) * | 2014-05-07 | 2016-03-02 | 山东博科生物产业有限公司 | A kind of homocysteine detection kit of strong interference immunity |
| CN103954774B (en) * | 2014-05-09 | 2016-02-03 | 山东博科生物产业有限公司 | A kind of stable homocysteine detection kit |
| CN104630324B (en) * | 2015-02-28 | 2016-11-09 | 北京爱必信生物技术有限公司 | Improved homocysteine detection reagent and method |
| CN105823744B (en) * | 2016-03-22 | 2020-04-03 | 江南大学 | Cysteine detection method and detection kit |
| CN106053830A (en) * | 2016-05-31 | 2016-10-26 | 安徽伊普诺康生物技术股份有限公司 | Kit for determining homocysteine and preparation method thereof |
-
2004
- 2004-03-08 CN CNB2004100167896A patent/CN100554948C/en not_active Expired - Lifetime
Non-Patent Citations (2)
| Title |
|---|
| Enzyme conversion immunoassay for determiningtotalhomocysteine in plasma or serum. Frank Frantzen, Arne Ludvig Faaren,Ingrid Alfheim.Clinical Chemistry,Vol.44 No.2. 1998 |
| Enzyme conversion immunoassay for determiningtotalhomocysteine in plasma or serum. Frank Frantzen, Arne Ludvig Faaren,Ingrid Alfheim.Clinical Chemistry,Vol.44 No.2. 1998 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN1560610A (en) | 2005-01-05 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104630324B (en) | Improved homocysteine detection reagent and method | |
| CN105181970B (en) | Stable kit for detecting homocysteine | |
| CN100554948C (en) | Homocysteine detection method and reagent thereof | |
| EP1644741B1 (en) | Methods and compositions for assaying homocysteine | |
| CN111944872A (en) | Reagent combination, reagent or kit for measuring creatinine content | |
| CN101498662A (en) | Reagent kit for monoamine oxidase MAO single-reagent measurement | |
| CN103558370A (en) | Carbon dioxide measurement kit | |
| CN102243227B (en) | Measuring method of asymmetric dimethylarginine concentration and measuring reagent thereof | |
| CN102095696A (en) | Homocysteine measuring method and reagent | |
| CN102154444B (en) | Method for measuring concentration of asymmetric dimethylarginine and diagnostic reagent kit | |
| CN109112181A (en) | A kind of 5`- activity of 5 '-nucleotidase measuring method and 5`- nucleotidase detection kit | |
| CN101514983A (en) | Reagent for determining serum total carnitine | |
| CN102140495B (en) | Method for testing dimethyl arginine dimethylamine hydrolytic enzyme and diagnostic reagent thereof | |
| JPWO2006030866A1 (en) | Method for quantitative determination of uric acid | |
| JPH04349898A (en) | Method and composition for determining d-glyceroaldehyde-3-phosphoric acid, inoganic phosphorus, or 1,3-diphosphoglyceric acid in high sensitivity | |
| CN115637265A (en) | Enzyme liquid protective agent for homocysteine circulating enzyme method detection and preparation method thereof | |
| CN101750355A (en) | Adenosine deaminase diagnostic reagent (kit) and adenosine deaminase activity concentration measuring method | |
| CN1757755B (en) | Determination method of creatnine content and reagent box for diagnosing creatnine | |
| CN117604068A (en) | Glycerol kinase activity determination method and determination kit | |
| CN101620142A (en) | Inorganic phosphorus (phosphate radical) diagnosis/determination kit and method for determining inorganic phosphorus (phosphate radical) concentration | |
| CN115343237A (en) | Kit and method for determining content of glucose-6-phosphate in biological sample | |
| JPS6028279B2 (en) | Reagent for measuring enzyme activity involved in purine metabolism | |
| Roberts et al. | Biomedical Science in Brief | |
| CN102081082A (en) | Determination method of galactose and galactose diagnosis/determination kit | |
| JP2001299389A (en) | Measuring method using enzyme reaction system comprising hydrolysis of adenosine-3-phosphoric acid, and measuring reagent to be used in the method |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| ASS | Succession or assignment of patent right |
Owner name: JIN JIANRUI Free format text: FORMER OWNER: WANG XUEZHONG Effective date: 20100531 |
|
| C41 | Transfer of patent application or patent right or utility model | ||
| COR | Change of bibliographic data |
Free format text: CORRECT: ADDRESS; FROM: 200125 ROOM 102, NO.14, LANE 2993, DONGFANG ROAD, PUDONG NEW AREA, SHANGHAI CITY TO: 100044 ROOM 319, BUILDING 1, SHIDAIMINGYUAN RESIDENTIAL QUATER, NO.45, XIZHIMEN NORTH STREET, BEIJING |
|
| TR01 | Transfer of patent right |
Effective date of registration: 20100531 Address after: 100044 Beijing City, Xizhimen No. 45 North Street District No. 1 era Mingyuan building room 319 Patentee after: Jin Jianrui Address before: 200125, room 14, No. 2993, Lane 102, Dongfang Road, Shanghai, Pudong New Area Patentee before: Wang Xuezhong |
|
| ASS | Succession or assignment of patent right |
Owner name: JIN JIANRUI Effective date: 20131115 Owner name: BEIJING ABZYMO BIOSCIENCES TECHNOLOGY CO., LTD. Free format text: FORMER OWNER: JIN JIANRUI Effective date: 20131115 |
|
| C41 | Transfer of patent application or patent right or utility model | ||
| COR | Change of bibliographic data |
Free format text: CORRECT: ADDRESS; FROM: 100044 HAIDIAN, BEIJING TO: 100176 DAXING, BEIJING |
|
| TR01 | Transfer of patent right |
Effective date of registration: 20131115 Address after: 100176, Beijing economic and Technological Development Zone, Hongda North Road, No. 12, block B, zone three, room, room two, 211-215 Patentee after: ABZYMO BIOSCIENCES Co.,Ltd. Patentee after: Jin Jianrui Address before: 100044 Beijing City, Xizhimen No. 45 North Street District No. 1 era Mingyuan building room 319 Patentee before: Jin Jianrui |
|
| ASS | Succession or assignment of patent right |
Owner name: KUNMING BEIAO BIOTECHNOLOGY CO., LTD. Free format text: FORMER OWNER: JIN JIANRUI Effective date: 20140603 |
|
| C41 | Transfer of patent application or patent right or utility model | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20140603 Address after: 100176, Beijing economic and Technological Development Zone, Hongda North Road, No. 12, innovation building, block B, zone 3, 210, Beijing Patentee after: ABZYMO BIOSCIENCES Co.,Ltd. Patentee after: Kunming times Biotechnology Co.,Ltd. Address before: 100176, Beijing economic and Technological Development Zone, Hongda North Road, No. 12, block B, zone three, room, room two, 211-215 Patentee before: ABZYMO BIOSCIENCES Co.,Ltd. Patentee before: Jin Jianrui |
|
| TR01 | Transfer of patent right |
Effective date of registration: 20180607 Address after: 225300 Taizhou, Jiangsu, China, west side of Tai Chi Road, China's medicine, building G59, 63 to three, west of Lu Jia Lu. Co-patentee after: Kunming times Biotechnology Co.,Ltd. Patentee after: JIANGSU DAJUN BIOTECHNOLOGY Co.,Ltd. Address before: 100176 Beijing city Beijing economic and Technological Development Zone Hongda North Road 12 innovation building B block 3 District 210 Co-patentee before: Kunming times Biotechnology Co.,Ltd. Patentee before: ABZYMO BIOSCIENCES Co.,Ltd. |
|
| TR01 | Transfer of patent right | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20200604 Address after: 225300 Jiangsu city of Taizhou province China Road West of Lu Jia Lu, Chengkou pharmaceutical G59 East Building 63, one to three layers of the West Co-patentee after: SHAN DONG CLINISCIENCES BIOTECH Co.,Ltd. Patentee after: JIANGSU DAJUN BIOTECHNOLOGY Co.,Ltd. Address before: 225300 Jiangsu city of Taizhou province China Road West of Lu Jia Lu, Chengkou pharmaceutical G59 East Building 63, one to three layers of the West Co-patentee before: Kunming times Biotechnology Co.,Ltd. Patentee before: JIANGSU DAJUN BIOTECHNOLOGY Co.,Ltd. |
|
| TR01 | Transfer of patent right | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20210824 Address after: Room 703-01, 7 / F, building 1, No. A5, Rongchang East Street, Beijing Economic and Technological Development Zone, Daxing District, Beijing 100101 Patentee after: BEIJING ANBAISHENG DIAGNOSIS TECHNOLOGY Co.,Ltd. Patentee after: SHAN DONG CLINISCIENCES BIOTECH Co.,Ltd. Address before: 225300 Taizhou, Jiangsu, China, west side of Tai Chi Road, China's medicine, building G59, 63 to three, west of Lu Jia Lu. Patentee before: JIANGSU DAJUN BIOTECHNOLOGY Co.,Ltd. Patentee before: SHAN DONG CLINISCIENCES BIOTECH Co.,Ltd. |
|
| TR01 | Transfer of patent right | ||
| IP01 | Partial invalidation of patent right |
Commission number: 4W114869 Conclusion of examination: Declare claims 1-2 of invention number 200410016789.6 invalid, and continue to maintain the validity of this patent on the basis of claims 3-6 Decision date of declaring invalidation: 20230327 Decision number of declaring invalidation: 561050 Denomination of invention: Determination method and reagents for homocysteine Granted publication date: 20091028 Patentee: Beijing Anbaisheng Diagnostic Technology Co.,Ltd.| Shandong Kelisen Biotechnology Co.,Ltd. |
|
| IP01 | Partial invalidation of patent right | ||
| CX01 | Expiry of patent term | ||
| CX01 | Expiry of patent term |
Granted publication date: 20091028 |