Fluorescent Determination of Creatine Phosphokinase Isoenzymes

Abstract

A high sensitive micro assay for Creatine phosphokinase (CPK) isoenzyme activity based on Bondar's method is described.
Three kinds of fraction of CPK isoenzyme, MM, MB and BB, in serum were separated by means of stepwise gradient liquid chromatography through a micro column contained DEAE Sepharose CL-6B (1.5×70mm) utilised three kinds of buffered sodium chloride solution (pH 7.5) that are 30mM for MM, 145mM for MB and 300mM for BB.
The separated fractions were determined its activity by spectrofluorometry (Ex 340nm, Em 460nm) of NADH generated from CPK reaction (Oliver-Rosalki s method).
It is able to measure the activity of CPK about 0.1mU/mleasily, so this method is applicable to a low activity of CPK isoenzyme such as BB and MB in the case of early myocardial infarction and a micro sample of new born baby.