DK156668B - Fremgangsmaade til kvantitativ bestemmelse af blodkoagulationsfaktor xii i humant plasma - Google Patents
Fremgangsmaade til kvantitativ bestemmelse af blodkoagulationsfaktor xii i humant plasma Download PDFInfo
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- DK156668B DK156668B DK484682A DK484682A DK156668B DK 156668 B DK156668 B DK 156668B DK 484682 A DK484682 A DK 484682A DK 484682 A DK484682 A DK 484682A DK 156668 B DK156668 B DK 156668B
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- ylamino
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- factor xii
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- 108010080865 Factor XII Proteins 0.000 title claims abstract description 21
- 102000000429 Factor XII Human genes 0.000 title claims abstract description 21
- 238000000034 method Methods 0.000 title claims abstract description 21
- 210000002381 plasma Anatomy 0.000 claims abstract description 26
- 108010071241 Factor XIIa Proteins 0.000 claims abstract description 23
- 239000012190 activator Substances 0.000 claims abstract description 6
- -1 4-hydroxycyclohexylmethyl group Chemical group 0.000 claims description 11
- 235000010469 Glycine max Nutrition 0.000 claims description 6
- 244000068988 Glycine max Species 0.000 claims description 6
- 238000006243 chemical reaction Methods 0.000 claims description 5
- 238000003776 cleavage reaction Methods 0.000 claims description 5
- 230000007017 scission Effects 0.000 claims description 5
- 239000002253 acid Substances 0.000 claims description 4
- 230000004913 activation Effects 0.000 claims description 4
- 239000005995 Aluminium silicate Substances 0.000 claims description 3
- 235000012211 aluminium silicate Nutrition 0.000 claims description 3
- 125000003277 amino group Chemical group 0.000 claims description 3
- 229910052500 inorganic mineral Inorganic materials 0.000 claims description 3
- NLYAJNPCOHFWQQ-UHFFFAOYSA-N kaolin Chemical compound O.O.O=[Al]O[Si](=O)O[Si](=O)O[Al]=O NLYAJNPCOHFWQQ-UHFFFAOYSA-N 0.000 claims description 3
- 239000011707 mineral Substances 0.000 claims description 3
- 150000007524 organic acids Chemical class 0.000 claims description 3
- 238000002360 preparation method Methods 0.000 claims description 3
- CFWRDBDJAOHXSH-SECBINFHSA-N 2-azaniumylethyl [(2r)-2,3-diacetyloxypropyl] phosphate Chemical compound CC(=O)OC[C@@H](OC(C)=O)COP(O)(=O)OCCN CFWRDBDJAOHXSH-SECBINFHSA-N 0.000 claims description 2
- 125000003143 4-hydroxybenzyl group Chemical group [H]C([*])([H])C1=C([H])C([H])=C(O[H])C([H])=C1[H] 0.000 claims description 2
- AFSDNFLWKVMVRB-UHFFFAOYSA-N Ellagic acid Chemical compound OC1=C(O)C(OC2=O)=C3C4=C2C=C(O)C(O)=C4OC(=O)C3=C1 AFSDNFLWKVMVRB-UHFFFAOYSA-N 0.000 claims description 2
- ATJXMQHAMYVHRX-CPCISQLKSA-N Ellagic acid Natural products OC1=C(O)[C@H]2OC(=O)c3cc(O)c(O)c4OC(=O)C(=C1)[C@H]2c34 ATJXMQHAMYVHRX-CPCISQLKSA-N 0.000 claims description 2
- 229920002079 Ellagic acid Polymers 0.000 claims description 2
- 125000000217 alkyl group Chemical group 0.000 claims description 2
- 125000001797 benzyl group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C([H])([H])* 0.000 claims description 2
- 239000007853 buffer solution Substances 0.000 claims description 2
- 125000004432 carbon atom Chemical group C* 0.000 claims description 2
- 125000004210 cyclohexylmethyl group Chemical group [H]C([H])(*)C1([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C1([H])[H] 0.000 claims description 2
- DEKPYXUDJRABNK-UHFFFAOYSA-N dimethyl 5-aminobenzene-1,3-dicarboxylate Chemical compound COC(=O)C1=CC(N)=CC(C(=O)OC)=C1 DEKPYXUDJRABNK-UHFFFAOYSA-N 0.000 claims description 2
- 229960002852 ellagic acid Drugs 0.000 claims description 2
- 235000004132 ellagic acid Nutrition 0.000 claims description 2
- 239000003112 inhibitor Substances 0.000 claims description 2
- 125000001160 methoxycarbonyl group Chemical group [H]C([H])([H])OC(*)=O 0.000 claims description 2
- FAARLWTXUUQFSN-UHFFFAOYSA-N methylellagic acid Natural products O1C(=O)C2=CC(O)=C(O)C3=C2C2=C1C(OC)=C(O)C=C2C(=O)O3 FAARLWTXUUQFSN-UHFFFAOYSA-N 0.000 claims description 2
- 230000002255 enzymatic effect Effects 0.000 abstract description 5
- 230000015271 coagulation Effects 0.000 description 8
- 238000005345 coagulation Methods 0.000 description 8
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 6
- 229940122618 Trypsin inhibitor Drugs 0.000 description 5
- 101710162629 Trypsin inhibitor Proteins 0.000 description 5
- 239000000203 mixture Substances 0.000 description 5
- 239000000758 substrate Substances 0.000 description 5
- 239000002753 trypsin inhibitor Substances 0.000 description 5
- 102000007469 Actins Human genes 0.000 description 3
- 108010085238 Actins Proteins 0.000 description 3
- KRKNYBCHXYNGOX-UHFFFAOYSA-K Citrate Chemical compound [O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O KRKNYBCHXYNGOX-UHFFFAOYSA-K 0.000 description 3
- 108090000790 Enzymes Proteins 0.000 description 3
- 102000004190 Enzymes Human genes 0.000 description 3
- MUBZPKHOEPUJKR-UHFFFAOYSA-N Oxalic acid Chemical compound OC(=O)C(O)=O MUBZPKHOEPUJKR-UHFFFAOYSA-N 0.000 description 3
- 239000007864 aqueous solution Substances 0.000 description 3
- 239000000872 buffer Substances 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
- 150000001875 compounds Chemical class 0.000 description 3
- 238000005259 measurement Methods 0.000 description 3
- 238000002156 mixing Methods 0.000 description 3
- TYMLOMAKGOJONV-UHFFFAOYSA-N 4-nitroaniline Chemical group NC1=CC=C([N+]([O-])=O)C=C1 TYMLOMAKGOJONV-UHFFFAOYSA-N 0.000 description 2
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 description 2
- 235000011054 acetic acid Nutrition 0.000 description 2
- 238000003556 assay Methods 0.000 description 2
- WPYMKLBDIGXBTP-UHFFFAOYSA-N benzoic acid Chemical compound OC(=O)C1=CC=CC=C1 WPYMKLBDIGXBTP-UHFFFAOYSA-N 0.000 description 2
- 238000011088 calibration curve Methods 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 230000002950 deficient Effects 0.000 description 2
- XBDQKXXYIPTUBI-UHFFFAOYSA-N dimethylselenoniopropionate Natural products CCC(O)=O XBDQKXXYIPTUBI-UHFFFAOYSA-N 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 238000011534 incubation Methods 0.000 description 2
- 230000002401 inhibitory effect Effects 0.000 description 2
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 2
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 2
- XNGIFLGASWRNHJ-UHFFFAOYSA-N phthalic acid Chemical compound OC(=O)C1=CC=CC=C1C(O)=O XNGIFLGASWRNHJ-UHFFFAOYSA-N 0.000 description 2
- 150000003839 salts Chemical class 0.000 description 2
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 description 1
- 239000005711 Benzoic acid Substances 0.000 description 1
- BHPQYMZQTOCNFJ-UHFFFAOYSA-N Calcium cation Chemical compound [Ca+2] BHPQYMZQTOCNFJ-UHFFFAOYSA-N 0.000 description 1
- 150000008574 D-amino acids Chemical group 0.000 description 1
- FEWJPZIEWOKRBE-JCYAYHJZSA-N Dextrotartaric acid Chemical compound OC(=O)[C@H](O)[C@@H](O)C(O)=O FEWJPZIEWOKRBE-JCYAYHJZSA-N 0.000 description 1
- 101100453790 Drosophila melanogaster Kebab gene Proteins 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 1
- CPELXLSAUQHCOX-UHFFFAOYSA-N Hydrogen bromide Chemical compound Br CPELXLSAUQHCOX-UHFFFAOYSA-N 0.000 description 1
- SNDPXSYFESPGGJ-UHFFFAOYSA-N L-norVal-OH Natural products CCCC(N)C(O)=O SNDPXSYFESPGGJ-UHFFFAOYSA-N 0.000 description 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 1
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 1
- FEWJPZIEWOKRBE-UHFFFAOYSA-N Tartaric acid Natural products [H+].[H+].[O-]C(=O)C(O)C(O)C([O-])=O FEWJPZIEWOKRBE-UHFFFAOYSA-N 0.000 description 1
- 208000007536 Thrombosis Diseases 0.000 description 1
- 150000007513 acids Chemical class 0.000 description 1
- YBCVMFKXIKNREZ-UHFFFAOYSA-N acoh acetic acid Chemical compound CC(O)=O.CC(O)=O YBCVMFKXIKNREZ-UHFFFAOYSA-N 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 125000003295 alanine group Chemical group N[C@@H](C)C(=O)* 0.000 description 1
- 125000000539 amino acid group Chemical group 0.000 description 1
- 235000010233 benzoic acid Nutrition 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 229910001424 calcium ion Inorganic materials 0.000 description 1
- 235000015165 citric acid Nutrition 0.000 description 1
- 239000000701 coagulant Substances 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 239000003527 fibrinolytic agent Substances 0.000 description 1
- 230000003480 fibrinolytic effect Effects 0.000 description 1
- 239000007850 fluorescent dye Substances 0.000 description 1
- 235000019253 formic acid Nutrition 0.000 description 1
- 208000031169 hemorrhagic disease Diseases 0.000 description 1
- RAXXELZNTBOGNW-UHFFFAOYSA-N imidazole Substances C1=CNC=N1 RAXXELZNTBOGNW-UHFFFAOYSA-N 0.000 description 1
- 230000002452 interceptive effect Effects 0.000 description 1
- 229950003188 isovaleryl diethylamide Drugs 0.000 description 1
- 235000015231 kebab Nutrition 0.000 description 1
- 239000004310 lactic acid Substances 0.000 description 1
- 235000014655 lactic acid Nutrition 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 235000006408 oxalic acid Nutrition 0.000 description 1
- 235000011007 phosphoric acid Nutrition 0.000 description 1
- 235000019260 propionic acid Nutrition 0.000 description 1
- IUVKMZGDUIUOCP-BTNSXGMBSA-N quinbolone Chemical compound O([C@H]1CC[C@H]2[C@H]3[C@@H]([C@]4(C=CC(=O)C=C4CC3)C)CC[C@@]21C)C1=CCCC1 IUVKMZGDUIUOCP-BTNSXGMBSA-N 0.000 description 1
- 229960000948 quinine Drugs 0.000 description 1
- 239000001509 sodium citrate Substances 0.000 description 1
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 description 1
- 238000002798 spectrophotometry method Methods 0.000 description 1
- 235000011149 sulphuric acid Nutrition 0.000 description 1
- 239000011975 tartaric acid Substances 0.000 description 1
- 235000002906 tartaric acid Nutrition 0.000 description 1
- YNJBWRMUSHSURL-UHFFFAOYSA-N trichloroacetic acid Chemical compound OC(=O)C(Cl)(Cl)Cl YNJBWRMUSHSURL-UHFFFAOYSA-N 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/56—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving blood clotting factors, e.g. involving thrombin, thromboplastin, fibrinogen
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
- C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
- C07K5/08—Tripeptides
- C07K5/0802—Tripeptides with the first amino acid being neutral
- C07K5/0804—Tripeptides with the first amino acid being neutral and aliphatic
- C07K5/0808—Tripeptides with the first amino acid being neutral and aliphatic the side chain containing 2 to 4 carbon atoms, e.g. Val, Ile, Leu
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2337/00—N-linked chromogens for determinations of peptidases and proteinases
- C12Q2337/10—Anilides
- C12Q2337/12—Para-Nitroanilides p-NA
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/90—Enzymes; Proenzymes
- G01N2333/914—Hydrolases (3)
- G01N2333/948—Hydrolases (3) acting on peptide bonds (3.4)
- G01N2333/95—Proteinases, i.e. endopeptidases (3.4.21-3.4.99)
- G01N2333/964—Proteinases, i.e. endopeptidases (3.4.21-3.4.99) derived from animal tissue
- G01N2333/96425—Proteinases, i.e. endopeptidases (3.4.21-3.4.99) derived from animal tissue from mammals
- G01N2333/96427—Proteinases, i.e. endopeptidases (3.4.21-3.4.99) derived from animal tissue from mammals in general
- G01N2333/9643—Proteinases, i.e. endopeptidases (3.4.21-3.4.99) derived from animal tissue from mammals in general with EC number
- G01N2333/96433—Serine endopeptidases (3.4.21)
- G01N2333/96441—Serine endopeptidases (3.4.21) with definite EC number
- G01N2333/96458—Factor XII (3.4.21.38)
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Zoology (AREA)
- Molecular Biology (AREA)
- Biophysics (AREA)
- Biochemistry (AREA)
- Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Microbiology (AREA)
- Immunology (AREA)
- Biotechnology (AREA)
- Analytical Chemistry (AREA)
- Physics & Mathematics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- Neurosurgery (AREA)
- Hematology (AREA)
- Medicinal Chemistry (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Peptides Or Proteins (AREA)
Description
DK 156668 B
Den foreliggende opfindelse angàr en fremgangsmâde til kvantitativ bestemmelse af koagulationsfaktor XII (ogsâ kaldeC Hageman-faktoren) i humant plasma.
Faktor XII er den ferste faktor, der aktiveres i den endogene blod» 5 koagulationskaskade. Da denne faktor ikke alene spiller en central rolle i koagulationssystemet, men ogsâ 1 kallikrein-kininsystemet og i det fibrinolytiske System, er det vigtigt at kunne bestemme denne faktor kvantitativt i blodplasma. Den for tiden mest anvendte bestem-melsesmetode bestâr 1, at en plasmapreve inkuberes med kaolin for at 10 aktivere Hageman-faktoren maksimalt, aktiveringsblandingen inkuberes med plasma, der mangler Hageman-faktoren, inkubationsblandingen efter et bestemt tidsrum recalcineres ved tilsætning af calciumioner, og koagulationstiden af den recalcinerede blanding mâles. Koagula-tionstiden er tilnarmelsesvis en funktion af koncentrationen af 15 Hageman-faktoren i testpreven (jfr. fx Oscar D. Ratnoff, "Thrombosis and Bleeding Disorders”, 1971, Georg Thieme Verlag, Stuttgart, side 215-218). Ulempen ved denne bestemmelsesmetode bestâr i, at man mâler en sterrelse, som er et résultat af en lang reaktionskaskade, og som derfor er afhangig af et stort antal reaktionsparametre. Bestemmelsen 20 vanskeliggeres yderligere ved, at der kraves vanskeligt tilgangeligt Hageman-faktor-manglende plasma til opstilling af kalibreringskurver.
I USA-patentskrift nr. 4.275.153 foreslâs det at anvende forbindelsen H-D-Phe-Pro-Arg-AIE (AIE - 5-aminoisophthalsyre-dimethylester) som substrat til bestemmelse af faktor Xlla i blodplasma. Faktor Xlla 25 spalter enzymatisk fra substratet en fluorescerende forbindelse, hvis mangde kan mâles fluorescensspektrofotometrisk. Den mængde faktor Xlla, der oprindeligt var til stede i blodplasmaet, kan bestemmes ud fra den iagttagne mangde. De resultater, der opnâs ved denne fremgangsmâde, er imidlertid ikke anvendelige, da forbindelsen H-D-Phe-30 Pro-Arg-AIE ikke alene spaltes af faktor Xlla, men ogsâ af kalli- krein, der ligeledes findes i blodplasma, oven i kebet ca. fem gange starkere end af faktor Xlla.
Det har nu vist sig at vare muligt direkte og kvantitativt at mâle den enzymatiske aktivitet af faktor Xlla, der er dannet ud fra faktor
DK 156668 B
2 XII, i blodplasma, idet der anvendes visse chromogene tripeptidde-rivater som spaltelige substrater.
Fremgangsmâden ifelge opfindelsen er ejendommelig ved, at plasmaet behandles med en aktivator for at omdanne den faktor XII, der fIndes 5 i plasma, til faktor Xlla, som omsættes med et tripeptidderivat med den almene formel H-D-NH-CH-CO-Gly-L-Arg-R1 R2 10 hvor R*- betegner en enzymatisk fraspaltelig chromogen eller fluorogen substitueret aminogruppe, og R2 betegner en ligekædet eller forgrenet alkylgruppe med 1-4 carbonatomer eller en benzyl-, p-hydroxybenzyl-, cyclohexylmethyl- eller 4-hydroxycyclohexylmethylgruppe, eller et sait deraf med en organlsk syre eller mineralsyre, og at mængden af 15 det ved hjslp af faktor Xlla fra tripeptidderivatet enzymatisk frigjorte farvede eller fluorescerende spaltningsprodukt R^H mâles fotometrisk, spektrofotometrisk eller fluorescensspektrofotometrisk.
Som aktivator kan der fx anvendes et préparât, som indeholder cepbalin og ellagsyre, eller kaolin. Aktiveringen af faktor XII 20 udferes hensigtsmæssigt ved ca. 0°C. Omsstningen af faktor Xlla med tripeptidderivatet udferes fortrinsvis i nerværelse af sojabonne-trypsininhibitor i et puffersystem med en pH-værdi pâ 7,4-8,0, fortrinsvis 7,7, og med en ionstyrke pâ 0,025-0,2, fortrinsvis 0,05.
Saltene af tripeptidderivaterne kan være salte med mineralsyrer, fx 25 HCl, HBr, H2SO4 eller H3PO4, eller med en organisk syre, fx myresyre, eddikesyre, propionsyre, mælkesyre, citronsyre, oxalsyre, vinsyre, benzoesyre, phthalsyre, trichloreddikesyre eller trifluoreddikesyre.
De i overensstemmelse med opfindelsen anvendte tripeptidderivater kan fremstilles pâ i og for sig kendt mâde.
30 Den ovenfor anferte almene formel omfatter fx de folgende enkelte tripeptidderivater:
DK 156668 B
3 2AcOH.H-D-Ala-Gly-Arg-pNA, 2AcOH.H-D-But-Gly-Arg-pNA, 2AcOH.H-D-Val-Gly-Arg-pNA, 2AcOH.H-D-Nval-Gly-Arg-pNA, 2AcOH.H-D-Leu-Gly-Arg-pNA, 2AcOH.H-D-Nleu-Gly-Arg-pNA, 2AcOH.H-D-Ile-Gly-Arg-pNA, 2AcOH.H-D-Phe-Gly-Arg-pNA, 5 2AcOH.H-D-Tyr-Gly-Arg-pNA, 2AcOH.H-D-CHA-Gly-Arg-pNA, 2AcOH.H-D-CHT-Gly-Arg-pNA.
AcOH - eddikesyre
Ala - alanyl
Arg - arginyl 10 But - a-aminobutyryl CHA - 3-cyclohexyl-alanyl CHT - 3-(4-hydroxycyclohexyl)-alanyl
Gly - glycyl
Ile - isoleucyl 15 Leu - leucyl
Nleu - norleucyl
Nval - norvalyl
Phe - phenylalanyl
Tyr - tyrosyl 20 Val - valyl pNa - paranitroanilid.
De to aminosyregrupper, der er knyttet til D-aminosyregruppen, har L-form.
De ovennævnte tripeptidderivater er i og for sig kendte forbindelser, 25 hvor p-nitrophenylaminogruppen kan erstattes med en anden chromogen eller fluorogen substitueret aminogruppe fx en l-carboxy-2-nitro-phenyl-5-ylamino-, l-sulfo-2-nitro-phen-5-ylami.no, ^-naphthylamino-, 4-methoxy-/3-naphthylamino-, 5-nitro-a-naphthylamino-, quinon-5-ylamino-, 8-nitro-quinon-5-ylamino-, 4-methyl-coumar-7-ylamino- eller 30 l,3-di(methoxycarbonyl)phen-5-ylaminogruppe (afledt af 5-amino-isophthalsyre-dimethylester).
Til udferelse af fremgangsmâden ifclge opfindelsen kan der gâs frem pâ felgende mâde.
4
DK 156 66 8 B
Det citratplasma, hvis indhold af faktor XII skal bestemmes, blandes med et vandigt præparat, som indeholder cephalin og eilagsyre, fx Actin® (fremstillet af DADE, Miami, USA) eller Cephotest® (frem-stillet af Nyegaard & Co. AS, Oslo, Norge), og blandingen inkuberes i 5 nogle fâ minutter, fortrinsvis 4 minutter, ved lav temperatur, fortrinsvis 0eC, for at omdanne faktor XII til faktor Xlla. (Citratplasma er blodplasma, til hvilket der er sat 3%'s vandig, isotonisk natrivuncitrat for at gore det ikke-koagulerende). Det er vigtigt at arbejde ved lave temperaturer for at undgâ dannelsen af enzymcom-10 plexer med plasmairihibitoren o^-makroglobulin, da disse complexer ganske vist enzymatisk ville kunne spalte de som substrat anvendte tripeptidderivater, men med ringere hastighed end de tilsvarende frie enzymer. En alikvotdel af den vundne aktiveringsblanding ssttes der-efter til et testsystem, der fremstilles ved blanding af en puffer, 15 fx tris-imidazolpuffer med en pH-værdi pâ 7,4-8,0, fortrinsvis 7,7, og en ionstyrke pâ 0,025-0,2, fortrinsvis 0,05, og som indeholder sojabonnetrypsininhibitor, med en vandig oplosning af et af de ovenfor definerede tripeptidderivater og en vandig oplosning af aktivatoren (Actin® eller Cephotest®) og prsinkubation i nogle 20 minutter ved den temperatur, ved hvilken spaltningen af tripep-tidderivatet skal mâles. Mængden af farvet eller fluorescerende spaltningsprodukt, der frisættes pr. tidserihed, mâles derefter ved fotometriske, spektrofotometriske eller fluorescensspektrofoto-metriske metoder. Nâr spaltningsproduktet er p-nitroanilin, foretages 25 mâlingen fortrinsvis ved 405 nm.
Den sojabonnetrypsininhibitor, der findes i testsystemet, har den funktion selektivt at hæmme det ved aktiveringen af faktor XII til faktor Xlla samtidigt aktiverede plasmakallikrein for at hindre dettes enzymatiske virkning pâ det som substrat anvendte tripep-30 tidderivat. Derimod har sojabonnetrypsininhibitoren ingen hæmmende virkning pâ faktor Xlla.
Fremgangsmâden ifolge opfindelsen har flere fordele fremfor den hidtil kendte bestemmelsesmetode, idet den muliggor en direkte be-stemmelse af faktor XII via den aktiverede faktor Xlla, hvis enzyma-35 tiske aktivitet kan fastsættes i et enkelt trin ved hjslp af de ovenfor anforte tripeptidderivater. I den hidtil kendte metode blev
DK 156668 B
5 aktiviteten af faktor Xlla malt indirekte ved hjslp af koagulati-onstiden. Koagulationen finder ferst sted, efter at den af faktor Xlla udleste fuldstændige endogene koagulationsproces er lebet til ende. For at fâ sammenlignelige mâlingsværdier er det nedvendigt at 5 anvende Hageman- faktor-manglende plasma for at bestemme indflydelsen af faktor Xlla pâ koagulationstiden og opstille kalibreringskurver til dette formai. Den hidtil kendte metode, hvis endelige mâlings-resultat ferst kunne fàs efter forlebet af komplicerede ind i hin-anden gribende enzymatiske processer, er behæftet med talrige 10 ukontrollerbare fejlkilder og er derfor unejagtig. Det er yderst vanskeligt at fâ et plasma, der fuldstsendigt mangler faktor XII. I kliniske laboratorier er der i dag tendens til at anvende automatiske analyseapparater, med hvilke der hurtigt og nejagtigt kan udferes analyser med et minimum af personale. Fremgangsmâden ifelge opfin-15 delsen egner sig fortræffeligt til sâdanne automatiske analyseapparater, hvorimod den hidtil kendte metode kun kan udferes semi-kvantitativt, manuelt og pâ tidsrevende mâde af uddannet laboratorie-personale.
Opfindelsen belyses nsrmere ved nedenstâende eksempel.
20 EKSEMPEL
Et testsystem, der bestâr af 0,75 ml tris-imidazolpuffer med en pH-værdi pâ 7,7 og en ionstyrke pâ 0,05, som indeholder 0,1 mg soja-bennetrypsininhibitor pr. ml, 0,25 ml 2 x 10'^ molær vandig oplesning af 2AcOH.H-D-Leu-Gly-Arg-pNA og 0,20 ml vandigt aktiveret cepha-25 loplastinreagens (Actin®, fremstillet af DADE, Miami, USA), præinku-bëres i 4 minutter ved 37eC. Præinkubatet tilsættes 0,05 ml af et inkubat, der er fremstillet ved blanding af 0,1 ml normalt citrat-plasma og 0,2 ml vandigt aktiveret cephaloplastinreagens og inku-bation i 4 minutter ved 0°C. Efter blanding af begge komponenter 30 mâles kontinuerligt mængden af det ved faktor Xlla frisette p-ni- troanilin spektrofotometrisk ved 405 nm i 5 minutter. Aktiviteten af faktor Xlla i enzymenheder pr. ml plasma beregnes ud fra foregelsen i optisk densitet pr. minut.
Claims (6)
1. Fremgangsmâde til kvantitativ bestenmelse af blodkoagulations-faktor XII i humant blodplasma, kendetegnet ved, at plasmaet behandles med en aktivator 10 for at omdanne faktor XII, der fIndes i plasmaet, til faktor Xlla, som omssttes med et tripeptidderivat med den almene formel H-D-NH-CH-CO-Gly-L-Arg-R1 R2 hvor rA betegner en enzymatisk fraspaltelig chromogen eller fluorogen 15 substitueret aminogruppe, og R2 betegner en llgeksdet eller forgrenet alkylgruppe med 1-4 carbonatomer eller en benzyl-, p-hydroxÿbenzyl-, cyclohexylmethyl- eller 4-hydroxycyclohexylmethylgruppe, eller et sait deraf med en organisk syre eller mineralsyre, og at mængden af det ved hjslp af faktor Xlla fra tripeptidderivatet enzymatisk 20 frigjorte farvede eller fluorescerende spaltningsprodukt R% mâles fotometrisk, spektrofotometrisk eller fluorescensspektrofotometrisk.
2. Fremgangsmâde ifelge krav 1, kendetegnet ved, at der som aktivator anvendes et préparât, der Indeholder cephalin og ellagsyre, eller kaolin.
3. Fremgangsmâde ifelge krav 1 og 2, kendetegnet ved, at omsætningen af faktor Xlla med tri-peptidderivatet udferes i nærværelse af sojabonnétrypsininhlbitor.
4. Fremgangsmâde ifelge krav 1-3, DK 156668 B kendetegnet ved, at omsstningen udferes i et puffersystem med en pH-værdi pâ 7,4-8,0, og med en ionstyrke pâ 0,025-0,2.
5. Fremgangsmâde ifelge krav 1, kendetegnet ved, at aktiveringen af faktor XII udferes ved 5 0°C.
6. Fremgangsmâde ifelge krav 1, kendetegnet ved, at R*- betegner en p-nitrophenylami.no-, 1-carboxy-2-nitro-phen-5-ylamino-, l-sulfo-2-nitro-phen-5-ylamino-, β-naphthylamino-, 4-methoxy-/?-naphthylamino-, 5-nitro-a-naphthylamino-, 10 quinon-5-ylamino-,· Ç-nitro-quinon-5-ylamino-, 4-methyl-coumar-7- ylamino- eller l,3-di(methoxycarbonyl)phen-5-ylaminogruppe (afledt af 5-amino-isophthalsyre-dimethylester).
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CH697281 | 1981-11-02 | ||
| CH697281 | 1981-11-02 |
Publications (3)
| Publication Number | Publication Date |
|---|---|
| DK484682A DK484682A (da) | 1983-05-03 |
| DK156668B true DK156668B (da) | 1989-09-18 |
| DK156668C DK156668C (da) | 1990-02-12 |
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Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| DK484682A DK156668C (da) | 1981-11-02 | 1982-11-01 | Fremgangsmaade til kvantitativ bestemmelse af blodkoagulationsfaktor xii i humant plasma |
Country Status (9)
| Country | Link |
|---|---|
| US (1) | US4598043A (da) |
| EP (1) | EP0078764B1 (da) |
| JP (1) | JPS5886100A (da) |
| AT (1) | ATE17262T1 (da) |
| CA (1) | CA1184837A (da) |
| DE (1) | DE3268329D1 (da) |
| DK (1) | DK156668C (da) |
| ES (1) | ES516987A0 (da) |
| NO (1) | NO823619L (da) |
Families Citing this family (18)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4882272A (en) * | 1986-10-01 | 1989-11-21 | Temple University - Of The Commonwealth System Of Higher Education | High molecular weight kininogen assay |
| JP3213832B2 (ja) * | 1993-08-06 | 2001-10-02 | 日本臓器製薬株式会社 | 被検物質の活性測定法 |
| JP3213831B2 (ja) * | 1993-08-06 | 2001-10-02 | 日本臓器製薬株式会社 | 被検物質の活性測定方法 |
| US6913900B2 (en) * | 2001-08-29 | 2005-07-05 | Nippon Zoki Pharmaceutical Co., Ltd. | Plasma prekallikrein activation and kallikrein production assay |
| GB0208989D0 (en) * | 2002-04-19 | 2002-05-29 | Amersham Biosciences Uk Ltd | Methods for measuring enzyme activity |
| JP2006510896A (ja) * | 2002-12-20 | 2006-03-30 | アクシス−シールド ダイアグノスティックス リミテッド | Ｘｉｉａ因子の変異体 |
| DE102005003145B4 (de) * | 2005-01-21 | 2006-10-19 | Dade Behring Marburg Gmbh | Stablies, chromogenes Flüssigreagenz und dessen Verwendung in gerinnungsdiagnostischen Tests |
| US20070166344A1 (en) * | 2006-01-18 | 2007-07-19 | Xin Qu | Non-leaching surface-active film compositions for microbial adhesion prevention |
| GB0607515D0 (en) * | 2006-04-13 | 2006-05-24 | Axis Shield Diagnostics Ltd | Anti-factor xlla therapy |
| EP2333555A1 (de) | 2009-12-09 | 2011-06-15 | Siemens Healthcare Diagnostics Products GmbH | Heterogener Gerinnungstest |
| EP2465941A1 (de) | 2010-12-20 | 2012-06-20 | Siemens Healthcare Diagnostics Products GmbH | Verfahren zur simultanen Bestimmung mehrerer Gerinnungsproteasen |
| US10473612B2 (en) | 2014-09-26 | 2019-11-12 | Abbott Point Of Care Inc. | Cartridge device identification for coagulation assays in fluid samples |
| EP3198280B1 (en) | 2014-09-26 | 2021-05-26 | Abbott Point Of Care, Inc. | Sensors for assaying coagulation in fluid samples |
| CN107110875A (zh) | 2014-09-26 | 2017-08-29 | 雅培医护站股份有限公司 | 用在凝固测定中的鞣花酸制剂 |
| CN106999932A (zh) | 2014-09-26 | 2017-08-01 | 雅培医护站股份有限公司 | 用于流体样本中的凝结测定的具有流体结的盒设备 |
| EP3197603A1 (en) | 2014-09-26 | 2017-08-02 | Abbott Point of Care Inc. | Cartridge device with segmented fluidics for assaying coagulation in fluid samples |
| EP4043879A1 (en) | 2014-09-26 | 2022-08-17 | Abbott Point Of Care Inc | Single channel cartridge device for coagulation assays in fluid samples |
| US10247741B2 (en) | 2014-09-26 | 2019-04-02 | Abbott Point Of Care Inc. | Microfabricated device with micro-environment sensors for assaying coagulation in fluid samples |
Family Cites Families (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| SE380258B (sv) * | 1972-05-02 | 1975-11-03 | Bofors Ab | Nya diagnostiskt verksamma substrat med hog specificitet till trypsin och andra enzymer av typen peptidyl-peptid-hydrolaser |
| SE380257B (sv) * | 1972-05-02 | 1975-11-03 | Bofors Ab | Nya diagnostiskt verksamma substrat med hog specificitet till trombin och andra proteolytiska enzymer av typen peptidyl-peptid-hydrolaser |
| SE407058B (sv) * | 1974-12-05 | 1979-03-12 | Kabi Ab | Nya kromogena enzymsubstrat for serinproteaser |
| US4169015A (en) * | 1975-07-11 | 1979-09-25 | Ab Kabi | Novel chromogenic thrombin substrates |
| SE407571B (sv) * | 1975-07-11 | 1979-04-02 | Kabi Ab | Nya kromogena enzymsubstrat for serinproteaser |
| US4137225A (en) * | 1975-07-11 | 1979-01-30 | Ab Kabi | Novel chromogenic enzyme substrates |
| CH634662A5 (de) * | 1976-05-28 | 1983-02-15 | Pentapharm Ag | Verwendung von tripeptidderivaten zur quantitativen bestimmung von plasminogen-aktivatoren. |
| US4302538A (en) * | 1978-03-27 | 1981-11-24 | Ortho Diagnostics Inc. | Buffer system in an anti-thrombin III test |
| US4275153A (en) * | 1978-08-03 | 1981-06-23 | American Hospital Supply Corporation | Analytical fluorogenic substrates for proteolytic enzymes |
| DE3061860D1 (en) * | 1979-04-24 | 1983-03-17 | Marcel Jozefonvicz | Process for the determination of proteases and antiproteases |
-
1982
- 1982-10-13 EP EP82810427A patent/EP0078764B1/de not_active Expired
- 1982-10-13 DE DE8282810427T patent/DE3268329D1/de not_active Expired
- 1982-10-13 AT AT82810427T patent/ATE17262T1/de not_active IP Right Cessation
- 1982-10-28 US US06/437,207 patent/US4598043A/en not_active Expired - Lifetime
- 1982-10-29 ES ES516987A patent/ES516987A0/es active Granted
- 1982-10-29 CA CA000414506A patent/CA1184837A/en not_active Expired
- 1982-11-01 DK DK484682A patent/DK156668C/da not_active IP Right Cessation
- 1982-11-01 JP JP57190895A patent/JPS5886100A/ja active Granted
- 1982-11-01 NO NO823619A patent/NO823619L/no unknown
Also Published As
| Publication number | Publication date |
|---|---|
| DK156668C (da) | 1990-02-12 |
| EP0078764A1 (de) | 1983-05-11 |
| NO823619L (no) | 1983-05-03 |
| ES8400603A1 (es) | 1983-10-16 |
| DK484682A (da) | 1983-05-03 |
| EP0078764B1 (de) | 1986-01-02 |
| JPS5886100A (ja) | 1983-05-23 |
| CA1184837A (en) | 1985-04-02 |
| ES516987A0 (es) | 1983-10-16 |
| JPH0346119B2 (da) | 1991-07-15 |
| ATE17262T1 (de) | 1986-01-15 |
| US4598043A (en) | 1986-07-01 |
| DE3268329D1 (en) | 1986-02-13 |
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