Kinetic colorimetric measurement of serum lactate dehydrogenase activity

Abstract

A kinetic colorimetric assay procedure is described for measuring lactate dehydrogenase activity, in which the reduction of NAD is coupled to the reduction of a tetrazolium salt, 2 p iodophenyl 3 p nitrophenyl 5 phenyl tetrazolium chloride (INT), with phenazine methosulfate serving as an intermediate electron carrier. The molar absorptivity of the formazan of INT was found to be 19.3 X 103 at 503 nm, which provides a threefold increase in sensitivity over the ultraviolet (340 nm) kinetic assay. The results correlate well with those obtained by 340 nm kinetic methods and the procedure does not require an ultraviolet spectrophotometer. The method is simple, rapid, and does not rely on secondary enzyme standards. It requires only two reagents, which are stable.