Highly sensitive enzyme immunoassay for human brain aldolase C

Abstract

A sensitive sandwich-type enzyme immunoassay for brain-type isozyme of human aldolase C 4 was developed using purified antibodies specific to the C subunit. The antibodies were raised in rabbits by injecting the purified aldolase C 4, and purified by means of immunoaffinity chromatography on a column of aldolase C 4-coupled Sepharose. 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 the minimum detection limit of aldolase C 4 was 3 pg/tube. The assay was specific to the C subunit of aldolase (aldolase C). It cross-reacted about 60% with aldolase AC 3, 30% with aldolase A 2C 2, and 4% with aldolase A 3C, but showed no cross-reactivity with aldolase A 4, the muscle-type isozyme. Coefficients of variation in within-run and between-run precision studies for serum aldolase C were ≤ 11%. Serum aldolase C levels in healthy adults of various ages (16–59 yr old) and both sexes ranged from 8.74–18.9 ng/ml. Immunoreactive aldolase C in the extracts of various human tissues was determined. It was distributed at high concentrations in the central nervous tissue and heart and at significant levels in liver, adrenal glands and testis. The assay of aldolase C in cerebrospinal fluid or serum by employing this sensitive immunoassay might be useful in the diagnosis of neurological disorders or acute myocardial damage.