A microassay for tetrazolium-reductase activity of polymorphonuclear leukocytes--comparison with a test-tube technique.
A micromethod for quantitative INT test is described to assay the activity of human peripheral blood PMNL. PMNL are obtained from a pellet after the separation of mononuclear cells. The test is performed in microtiter plates with 96 wells. The estimation of the activity is based on the colour change following the INT reduction to formazane. As a stimulating agent starch (rice species--Amylum oryzae) or zymosan is used. Absorbance alternations are measured using a microplate reader (e.g. MR 580, Dynatech). Conditions necessary for standard results (i.e. PMNL concentration, type and concentration of stimulating agent, incubation time, sedimentation time for PMNL isolation) were verified experimentally. Blood samples of 60 blood donors were simultaneously tested by the micromethod and the test-tube method. The statistical analysis (F-test of homogeneity of variances, t-test with paired observations, correlation coefficient of tightness of dependence) proved that the results of the micromethod were comparable to the test-tube assay. The advantages of the micromethod are as follows: low blood consumption, possibility to perform more parallel examinations of the same sample, and to test more samples at a time, standard and highly reproducible results, saving of reagents, reduction of labour time, possibility of some further automation.
D000328:Adult; D001782:Blood Donors; D003124:Colorimetry / Q000379:methods; D006801:Humans; D007202:Indicators and Reagents; D007700:Kinetics; D008832:Microchemistry; D009246:NADH Tetrazolium Reductase / Q000097:blood*; D009247:NADH, NADPH Oxidoreductases / Q000097:blood*; D009504:Neutrophils / Q000201:enzymology*
D007202:Indicators and Reagents; D009246:NADH Tetrazolium Reductase; D009247:NADH, NADPH Oxidoreductases