Highly sensitive enzyme immunoassay for human creatine kinase BB isozyme

Area of Science:

• Biochemistry
• Immunology
• Clinical Diagnostics

Background:

• Creatine kinase isoenzymes, particularly CK-BB, are important biomarkers.
• Accurate measurement of CK-BB is crucial for diagnosing neurological and other disorders.
• Existing methods may lack sensitivity or specificity for CK-BB detection.

Purpose of the Study:

• To develop a highly sensitive and specific sandwich-type enzyme immunoassay for human creatine kinase isoenzyme BB (CK-BB).
• To validate the assay's performance characteristics, including sensitivity, specificity, and precision.
• To determine normal ranges of CK-BB in serum and cerebrospinal fluid (CSF) and assess its tissue distribution.

Main Methods:

• Development of a sandwich enzyme immunoassay using specific antibodies against the CK-B subunit.
• Immobilization of antibody F(ab')2 fragments on polystyrene balls and Fab' fragments labeled with beta-D-galactosidase.
• Measurement of CK-BB in human serum, cerebrospinal fluid, and tissue extracts.

Main Results:

• The developed assay demonstrated high sensitivity, measuring as low as 1 pg of CK-BB.
• The assay showed high specificity for the CK-B subunit, with minimal cross-reactivity with CK-MB and no cross-reactivity with CK-MM or enolase.
• Precision studies yielded coefficients of variation less than 8% for serum CK-B.
• Normal serum CK-BB levels were established for adults and children, with higher concentrations observed in children.
• CK-BB levels in CSF of patients with non-neuronal disorders were determined.
• Tissue analysis confirmed high CK-BB concentrations in the central nervous system and other specific organs.

Conclusions:

• The developed enzyme immunoassay is a sensitive, specific, and precise method for quantifying human CK-BB.
• The assay provides valuable data on CK-BB levels in various biological fluids and tissues.
• This method can aid in the diagnosis and understanding of conditions involving CK-BB.