A modified, optimized kinetic photometric assay for the determination of blood coagulation factor XIII activity in plasma

Background: Blood coagulation factor XIII (FXIII) is a zymogen that is tran sformed into an active transglutaminase by thrombin and Ca2+. FXIII plays a n essential role in fibrin stabilization and in the protection of fibrin fr om proteolytic degradation. No convenient method has been available for the measurement of FXIII activity in plasma. The aim of the present study was to improve and optimize a kinetic photometric FXIII assay originally developed in our laboratory. Methods: In the assay, FXIII was activated by thrombin and Ca2+. Fibrin polymerization was prevent ed by an inhibitory tetrapeptide. Glycine-ethyl ester and a glutamine resid ue of a synthetic dodecapeptide served as acyl acceptor and acyl donor tran sglutaminase substrates, respectively. The amount of ammonia released durin g the reaction was monitored using glutamate dehydrogenase and NADPH. Results: The use of a new glutamine substrate and optimization of activator and substrate concentrations increased sensitivity. Substitution of NADPH for NADH and introduction of an appropriate blank eliminated systemic overe stimation of FXIII activity. The recovery of FXIII was 96%, the assay was l inear up to 470 U/L, the detection limit was 1 U/L, and the imprecision (CV ) was <8% even at very low FXIII activities. A reference interval of 108-22 4 U/L (69-143%) was established. The results correlated well with results o btained by an immunoassay specific for plasma FXIII: Conclusions: The optimized FXIII assay is a simple, rapid method for the di agnosis of inherited or acquired FXIII deficiencies and increased FXIII con centrations. It can be easily adapted to clinical chemistry analyzers. (C) 2000 American Association for Clinical Chemistry.