KineticDeterminationof Serum Haptoglobin witha CentrifugalAnalyzer

We describe a method for determining haptoglobin with a centrifugal analyzer that is based on haptoglobin combining stoichiometrically with hemoglobin to form a complex that has peroxidase-like activity proportional to the quantity of haptoglobin present. Under assay conditions, unbound hemoglobin exhibits only a small fraction of the total peroxidase activity. Activity is measured colorimetrically at 405 nm after reaction with o-dianisidine and ethyl hydrogen peroxide. The procedure is standardized by saturating a known amount of hemoglobin with a serum whose hemoglobin binding capacity exceeds the amount of hemoglobin in the assay system. The mean and mean within-run precision of our method, determined by performing 17 replicate assays of both a pooled normal serum and a 10-fold dilution of the serum, was 1.13 g/liter (CV, 2.9%), and 106 mg/liter (CV, 5.8%), respectively. The 95 percentile estimate of the normal range by our method is 0.45-1.85 g/liter hemoglobin binding capacity. When results by our automated method were compared to those by a manual method [Scand. J. Gun. Lab. Invest. 18, 80(1965)], the slope of the unweighted linear leastsquares regression line was .970, the y-intercept 26 mg/liter, and the correlation coefficient .995.