An Automated Fluorometric Method for Determination of Lactic Dehydrogenase in Serum

A simple automated method for the determination of lactic dehydrogenase (LDH) activity in serum by the forward lactate- pyruvate reaction is presented. Enzyme activity is determined by measuringthe fluorescence of DPNH formed after a fixed incubation time. Detailed studies of various parameters of the reaction, reproduci-bility, and recovery data, and a comparisonwith two manual methods for LDII are presented. TE1HE DETERMINATION of serum lactic dehydrogenase activity (LDH) has proved to he important in the diagnosis of myocardial infarction (1-5) and helpful in the diagnosis and study of many diverse conditions, most of which have in common necrosis of tissue (1, 6). The increased demand for this assay in our laboratory, particularly as part of the analysis done in every suspected case of myocardial infarction, prompted us to attempt to develop a simple reliable method applicable to automatic analysis. Lactic dehydrogenase is an enzyme that reversibly catalyzes the con-version of lactate to pyruvate, with the simultaneous i’eduction of diphosphopyridine nucleotide (DPN) to reduced diphosphopyridiiie iiucleotide (DPNH). LDH Lactate + DPN Pyruvate + DPMI The rate of this reaction is proportional to the activity of LDH present (7, 8). LDH activity may be assayed either by measuring the change in the DPN-DPNH system, or by measuring the change in the lactate-pyruvate system. Many manual methods which measure the DPN-DPNH system have been described. These depend on the differences in optical properties of T)PN and DPNH: (1) I)PNH shows maxi