DISCONTINUED
Intended Use:
The Prothrombin kit is a chromogenic assay for the quantitative determination of Prothrombin associated with the G20210A mutation in human citrated plasma, using a manual or automated protocol. Research use only .
Summary:
Congenital or acquired Prothrombin deficiency is a risk factor for a bleeding disorder. Elevated prothrombin concentration could be associated with an increased thrombotic risk.
Assay Principle:
Prothrombin is measured following its specific activation with Ecarin, an enzyme extracted from snake venom (Echis Carinatus). Meizothrombin is formed and then it specifically cleaves the specific substrate SIIa-01, releasing para-nitroaniline (pNA), which color is measured at 405nm. There is a direct relationship between color development and Prothrombin activity in the tested plasma.
Reagents - Storage & Stability:
- R1: Reagent 1: 4 vials containing about 2 U of Ecarin. Highly purified enzyme, extracted from the Echis carinatus snake venom, lyophilized in the presence of a fibrin polymerization inhibitor, and stabilized; Ecarin can specifically activate prothrombin into meizothrombin: Reconstitute each vial with exactly 2.5 ml of distilled water; homogeneize the content. Let the reagent to stabilise for 30 min at Room Temperature, before use, while shaking from time to time. Homogeneize before use. Stability of reconstituted Ecarin, kept in its original vial: 1 week at 2-8°C or 3 days at Room Temperature (18-25°C). Do not freeze.
- R2: Reagent 2: 4 vials containing about 5 mg of Thrombin specific Chromogenic substrate (SIIa-01), lyophilized. Reconstitute each vial with 2.5 ml of distilled water; homogenize the content. Incubate at Room Temperature (18-25°C) for 30 min, before use, while shaking from time to time. Homogeneize before use. Stability of restored substrate, kept in its original vial: 1 month at 2-8°C or 3 days at Room Temperature (18-25°C). Do not freeze.
- R3: Reagent 3: 4 vials containing about 5 ml of TBSA buffer Ten fold concentrated "10xconc.". Contains BSA and sodium azide. Shake the vial and dilute the amount required 1:10 in distilled water (the 5 ml contained in the vial allow preparing 50ml of ready to use buffer). The buffer must be stored at 2-8°C in its original vial and used within 4 weeks following opening. The diluted buffer must be used within 7 days, when protected from any contamination and stored at 2-8°C.