Enzyme-linked immunosorbent assay (ELISA) for the measurement of small quantities of alpha 2-macroglobulin.

A simple, sensitive and precise enzyme-linked immunosorbent assay for the quantitation of alpha 2-macroglobulin (alpha 2M) in supernatants of cell cultures was constructed. All reagents apart from the alpha 2M standard were commercially available. The assay range was 2.0-500 micrograms/l. The intra-assay coefficient of variation (CV%) was 4.6%, and the imprecision between runs was 8.9% at 10 micrograms/l and 9.0% at 110 micrograms/l. Recovery of alpha 2M, added to cell culture medium free of serum, was 97.5 +/- 7.2% (mean +/- SD) and the recovery of alpha 2M added to pooled human serum was 101 +/- 6.0%. There was no significant difference between the recovery of alpha 2M-standard and alpha 2M-trypsin complexes, whereas the dose response of a commercial alpha 2M-standard was lower than expected (81.1 +/- 7.5%), indicating a lower purity and/or conformational changes in the epitopes of this reagent. As expected, supernatants of mononuclear lymphocyte cultures enriched in monocytes contained significantly higher concentrations of alpha 2M than supernatants of cell cultures depleted in monocytes. Our results indicate that the ELISA method could be useful tool in the study of the alpha 2M turnover in all cell cultures in vitro.