OPTIMIZATION OF A NEW CONTINUOUS UV ASSAY FOR THE DETERMINATION OF BLOOD-COAGULATION FACTOR-XIII ACTIVITY IN HUMAN PLASMA

The new photometric assay described by Fickenscher et al. (Thromb. Haemostas. 65 (1991) 535-540) for the determination of factor XIII facilitates the diagnosis of factor XIII deficiency. In spite of easy handling, this test should be used critically. Patients with hyperfibrinogenaemia showed factor XIII activities of less than 20%, whereas with an optimized assay we found normal factor XIII values. Also, the use of a fixed period of incubation for the analysis is questionable, because the period of constant reaction rate occurs earlier and is shorter with high factor XIII activities and later and longer with low factor XIII activities. A linear relation between factor XIII activity and signal only exists up to 80% of activity. In some plasma samples from patients with hyperfibrinogenaemia the factor XIII determination actually shows decreased values for factor XIII. During the reaction, a fibrin clot is formed. The resulting turbidity simulates an increase in absorbance so that NADH consumption is apparently decreased. In six patients with hyperfibrinogenaemia (8.1 -9.4 g/1), a factor XIII activity of 26 U/I or less was determined. Using 50 mul instead of 100 mul sample volume, 50% (3/6) of the patients showed a normal factor XIII activity (80-96 U/l), whereas 50% (3/6) values of 6 - 15 U/l were found. In our modified assay we measured normal factor XIII activities (72 - 151 U/l) in all 6 patients. The procedure is optimized by reducing the sample volume from 100 mul to 50 pl. The concentration of the fibrin aggregation inhibitor is raised from 0.5 g/l to 1.0 g/l in the test solution, and the period of constant reaction rate is used for the determination. Enzyme activity is expressed in U/l (37-degrees-C). Optimized in this way, the assay shows a linear relationship between the decrease of absorbance and factor XIII activity up to more than 128 U/l, corresponding to 140%. Plasma samples with high concentrations of fibrinogen can be analysed without problems. Precision was 2.3% in series and 2.8% from day to day, using a control plasma with 81 U/l; for a control plasma with 35 U/1, the respective values were 4.9% and 5.8%. In apparently healthy 100 persons a reference interval of factor XIII activity was established from 66 to 142 U/l or 73 to 155%.