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Standardization of creatine kinase-MB (CK-MB) mass assays: the use of recombinant CK-MB as a reference material
R H Christenson1, H Vaidya, Y Landt
1Department of Pathology, University of Maryland School of Medicine, Baltimore, MD 21201, USA. rchriste@umaryland.edu
Background:
The AACC assembled a committee to identify and validate a standard creatine kinase MB isoenzyme (CK-MB) material to improve the comparability of CK-MB mass assays.
Methods:
Three protocols were used. In protocol I, various CK-MB materials prepared in different matrices were screened as candidate standards. In protocol II, participating manufacturers calibrated their systems with concentrates of human heart CK-MB and then tested 20 patient samples to evaluate calibration bias. In protocol III, participating manufacturers calibrated their immunoassay systems using recombinant CK-MB2 (rCK-MB2) diluted into their respective sample diluents and measured 50 samples.
Results:
Candidate materials showed high recovery in stripped human serum, but bias improved only from 59% to 38%. These data led to the use of human heart CK-MB diluted in each manufacturer's sample diluent. This strategy reduced bias from 31% to 15%. Because human heart CK-MB is difficult to provide, a lyophilized source of CK-MB2 was identified. rCK-MB2 was shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, reversed-phase HPLC, intrinsic protein fluorescence, circular dichroism, agarose gel electrophoresis, immunoreactivity studies, high and low temperature stability, and reconstituted stability to be equivalent to human heart CK-MB. Calibration of immunoassay systems with rCK-MB2 added into each respective manufacturer's sample diluent showed a 13% between-manufacturer bias.
Conclusion:
Lyophilized rCK-MB2 was determined suitable for use as a reference material for CK-MB mass assays.