Journal oflmmunologicalMethods, 87 (1986) 229-237 229 Elsevier JIM03815 Quantitation of rheumatoid factors by enzyme immunoassay using biotinylated human IgG M. Zrein, G. De Marcillac and M.H.V. Van Regenmortel lnstitut de Biologie MolOculaire et Cellulaire du CNRS, 15, rue Descartes, 67000 Strasbourg, France (Received 13 September 1985, accepted I November 1985) An enzyme-linked immunosorbent assay (ELISA) using biotinylated human IgG for the detection of rheumatoid factor (RF) in human sera has been developed. This assay avoids some of the problems encountered with earlier ELISA methods used for RF detection. Accurate RF titers of human sera were determined using a simple computer program developed for curve fitting of ELISA data. Key words: Rheumatoid factor; ELISA; Biotin Introduction Rheumatoid factors (RF) are autoantibodies that react with epitopes situated on the Fc portion of mammalian IgG. The presence of RF in human serum is indicative of rheumatoid arthritis al- though RF are also found in patients with various infections and chronic inflammatory conditions. Although RF occur in all major immunoglobulin classes, RF of the IgM class usually predominate. The classical methods for detecting RF are the hemagglutination method of Waaler (1940) and Rose et al. (1948) and the latex test of Singer and Plotz (1956) both of which mainly measure IgM RF. In recent years, various solid-phase immunoas- says for the measurement of RF have been de- scribed. In all these tests, RF are bound to a solid phase precoated with IgG and are then revealed by the addition of reagents such as radiolabelled anti- body to human IgM (Knez and Reimer, 1977), enzyme-labelled human IgG (Maiolini et al., 1978; Togun and Brenchley, 1983) or enzyme-labelled antibody to human immunoglobulin classes (Vejtorp et al., 1979; Gripenberg et al., 1979; Halbert et al., 1980). Although the sensitivity of these solid-phase immunoassays is entirely adequate, they require the use of labelled reagents (for instance anti-hu- man IgM) that do not react with the human IgG used for trapping the RF on the solid phase. Since most commercial reagents (conventional antisera) specific for human/~-chain cross-react with human IgG, it is necessary either to cross-absorb them (Halbert et al., 1980) or to dilute them sufficiently to avoid the cross-reaction (Vejtorp et al., 1979). Alternatively, highly specific monoclonal antibod- ies can also be used (Teitsson and Valdimarsson, 1984). For laboratories engaged in routine diagno- sis the need for such precautions increases the cost and limits the usefulness of the test. In this report, we describe an enzyme-linked immunosorbent assay (ELISA) in which enzyme- labelled antibody used for revealing RF is re- placed by biotin-labelled human IgG followed with enzyme-labelled avidin. This procedure avoids the problems that arise when the labelled antibody used to detect the RF cross-reacts with the IgG 0022-1759/86/$03.50 © 1986 Elsevier Science Publishers B.V. (Biomedical Division)