A convenient and sensitive sequential sandwich colorimetric ELISA test was established for quantitating IL-6 in culture supernatants or in serum. Immunopurified HRP-labelled rabbit Fab' fragment was used as the tracer and IgG-coated microtiter plate as the capture antibody. The limit of detection was as low as 10 attomoles of analyte (2.5 pg/ml). Unglycosylated recombinant IL-6 and the natural glycosylated cytokine were recognized equally. In addition, IL-6 measurements were unaffected by the presence of various cytokines and assay sensitivity was only slightly reduced in the presence of undiluted serum samples. The technique was applied to the study of in vitro IL-6 production from activated monocytes and to the in vivo determination of IL-6 in various pathological states.