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Measurement of Factor V Activity in Human Plasma Using a Microplate Coagulation Assay
Published on: September 9, 2012
Quantification of coagulation factor XIII activity by a thio-NADH based assay using factor XIII immuno-depleted
Andreas Kappel1, Sina Stephan, Gerlinde Christ
1Siemens Healthcare Diagnostics Products GmbH, Marburg, Germany.
Background:
Accurate determination of factor XIII (FXIII) activity is crucial for replacement therapy. FXIII activity is typically determined using a coupled enzymatic reaction that measures nicotinamide adenine dinucleotide hydride (NADH) consumption at 340 nm.
Methods:
Here, we describe the development of a prototype for a novel FXIII activity assay for detection at 405 nm by replacing NADH with thio-NADH, and the application of FXIII immuno-depleted plasma as a diluent for calibration.
Results:
Performance data show up to two-fold lower susceptibility of the prototype assay to interferences from hemolyzed, icteric, and lipemic samples when compared to a NADH assay format. In addition, the use of FXIII immuno-depleted plasma as diluent for calibration improved recovery almost two-fold in the lower measurement range. The novel prototype assay correlates well with a conventional assay (r=0.98, y=0.99·x+2.17% FXIII, n=173).
Conclusions:
The described prototype assay has the potential to (a) increase trueness of measurement of low levels of FXIII, (b) improve robustness due to reduction from interferences, and (c) can be used on a broad range of coagulation instruments due to its detection at 405 nm.
