title: Standardization of creatine kinase-MB (CK-MB) mass assays: the use of recombinant CK-MB as a reference material.
abstract: 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.
  
  
  CONCLUSIONS
  Lyophilized rCK-MB2 was determined suitable for use as a reference material for CK-MB mass assays.
authors.affiliation: Department of Pathology, University of Maryland School of Medicine, Baltimore, MD 21201, USA. rchriste@umaryland.edu
authors.forename: R H
authors.initials: RH
authors.lastname: Christenson
authors.forename: H
authors.initials: H
authors.lastname: Vaidya
authors.forename: Y
authors.initials: Y
authors.lastname: Landt
authors.forename: R S
authors.initials: RS
authors.lastname: Bauer
authors.forename: S F
authors.initials: SF
authors.lastname: Green
authors.forename: F A
authors.initials: FA
authors.lastname: Apple
authors.forename: A
authors.initials: A
authors.lastname: Jacob
authors.forename: G R
authors.initials: GR
authors.lastname: Magneson
authors.forename: S
authors.initials: S
authors.lastname: Nag
authors.forename: A H
authors.initials: AH
authors.lastname: Wu
authors.forename: H M
authors.initials: HM
authors.lastname: Azzazy
mesh_terms: D002138:Calibration; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D003402:Creatine Kinase / Q000032:analysis / Q000302:isolation & purification / Q000592:standards*; D004587:Electrophoresis, Agar Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006801:Humans; D007527:Isoenzymes; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000592:standards; D012015:Reference Standards; D015203:Reproducibility of Results
chemical_list: D007527:Isoenzymes; D011994:Recombinant Proteins; D003402:Creatine Kinase