Abstract
A conventional method for the estimation of plasma α2-plasmin inhibitor (α2-PI) was proposed upon introduction of a novel chromogenic substrate, yet procedures were still incommoded as routine assay method. We present a improved procedure so as to put this method in running order.
Procedure: Citrated plasma to be assayed is diluted first 101 folds with Tris-HCl buffer (pH 7.5) and 0.2ml of aliquot is incubated for 10 minutes at 37°C with 0.05ml of anti α2-macroglobulin (anti α2M) 0.1ml of plasmin solution is then added and stand for 10 minutes at the same temperature, followed by the addition of substrate solution (Tos-Gly-Pro-Lys-pNA) and incubate whole reaction mixture for further 10 minutes. Reaction is stopped by adding 2.5ml of acetic acid (10%) and read absorbancy at 405nm.
Results: Plasmin inhibiting activity of α2-macroglobulin in specimen sera appears to be completely blocked by the addition of anti α2M under the conditions provided. The fact that the estimated values with specimen sera are apparently the same as those of reference sera deprived of α2-macroglobulin by affinity chromatography, is revealing of the specificity of this method for the α2-PI assay. This assay method shows linearity up to 125% and the correlation coefficient of this method vs. the immunological method is 0.97.
