Clinical Chemistry, Vol 34, 575-581, Copyright © 1988 by American Association for Clinical Chemistry

Semi-automated direct colorimetric measurement of creatine kinase isoenzyme MB activity after extraction from serum by use of a CK-MB- specific monoclonal antibody

Y Landt, HC Vaidya, SE Porter, K Whalen, A McClellan, C Amyx, CA Parvin, G Kessler, MH Nahm and DN Dietzler
Department of Pathology, Washington University School of Medicine, St. Louis, MO 63110.

This semi-automated colorimetric assay for the MB isoenzyme of creatine kinase (EC 2.7.3.2) is based on a monoclonal antibody ("Conan-MB") specific for this isoenzyme and is a modification of a previously published method (Vaidya et al., Clin Chem 1986;32:657-63). A 0.64-cm bead coated with 2 to 3 micrograms of antibody is incubated with 100 microL of serum and 10 microL of 0.2 mol/L beta-mercaptoethanol for 1 h at room temperature, to extract CK-MB. The beads are washed with de- ionized water and incubated with CK substrate for 45 min at 37 degrees C. A solution containing trans-1,2-diaminocyclohexane-N,N,N', N'- tetraacetic acid, p-iodonitrotetrazolium violet, and diaphorase is added and the resulting colored product is measured at 492 nm. The standard curve is linear to 200 U of CK-MB per liter, and analytical recovery is 97-113%. Total assay CV for low (9.7 U/L) and high (50.7 U/L) quality-control materials was 14.1% (n = 1878) and 11.6% (n = 1842), respectively. CK-MB activity correlated well (r = 0.978, n = 226) with CK-MB measured by a two-site mass immunoassay, and 99.4% of samples with CK-MB greater than or equal to 12 U/L (n = 347) were verified by electrophoresis on agarose.