Highly sensitive enzyme immunoassay for human creatine kinase BB isozyme

Abstract

A sensitive sandwich-type enzyme immunoassay method for measurement of brain-type isozyme of human creatine kinase (CK-BB) was developed using purified antibodies specific to the B subunit. The assay system consisted of polystyrene balls with immobilized antibody F(ab') 2 fragments and the same antibody Fab' fragments labelled with β-D-galactosidase from Escherichia coli. The assay was highly sensitive and 1 pg of CK-BB was measurable. The assay was specific to the B subunit of creatine kinase (CK-B), and it cross-reacted about 25% with CK-MB, the heart-type isozyme. However, the assay showed no cross-reactivity with CK-MM, the muscle type-isozyme or with neuron-specific αα enolase. Coefficients of variation in withinrun and between-run precision studies for serum CK-B were < 8%. Serum CK-B levels in healthy adults of various ages (16–59 yr old) ranged from 0.25–1.44 ng/ml, whereas the CK-B concentrations in children (< 10 yr old) were relatively high, ranging from 1.3–7.4 ng/ml. The CK-B levels in the cerebrospinal fluids (CSF) could be determined by the present method, and they ranged from 0.10–0.76 ng/ml in the samples from patients with non-neuronal disorders. Determination of immunoreactive CK-B in the extracts of various human tissues confirmed previous reports that CK-B was distributed at high concentrations in the central nervous tissue, prostate, uterus, bladder, gastrointestinal tract and heart muscle.