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  • Quantification of prothrombin in human plasma amplified by autocatalytic reaction.

Quantification of prothrombin in human plasma amplified by autocatalytic reaction.

Analytical chemistry (2012-02-14)
Ana Virel, Laura Saa, Valeri Pavlov
ABSTRACT

By site directed mutagenesis, we have produced recombinant mutants of human and mouse prethrombin-2 which are able to convert themselves autocatalytically into α-thrombin. We also have created a new method to amplify the signal of bioanalytical assays based on the autocatalytic activation of these mutated proenzymes. The activation of the mutants by active α-thrombin triggers an autocatalytic reaction which leads to more active thrombin resulting in the amplification of the readout signal. Addition of mutated mouse prethrombin-2 into the conventional assay for prothrombin level in human plasma, employing ecarin and the fluorogenic substrate, resulted in improvement of the detection limit by 2 orders of magnitude.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Rhodamine 110 chloride, suitable for fluorescence, BioReagent, ≥99.0% (UV)
Sigma-Aldrich
Rhodamine 110 chloride, Dye content ≥75 %