Human fibroblast-derived molecules as antigens in enzyme-linked immunosorbent assay for coeliac disease-specific IgA.

We have recently shown that cultured human fibroblasts synthesize and secrete protein molecules that bind to IgA-class anti-reticulin and anti-endomysium antibodies but not to anti-gliadin antibodies in coeliac disease patient sera. In the present report, we describe a reproducible method for purification of these antigen molecules from fibroblast culture medium. Using reversed-phase chromatography as the final purification step, four different protein molecules reacting with coeliac disease patient sera IgA were obtained. In enzyme-linked immunosorbent assay (ELISA) for coeliac disease-specific IgA, a mixture of 0.5 microgram of the four reversed-phase-separated molecules was used as antigen. The optical density values in ELISA of the sera from newly diagnosed coeliac disease patients (n = 34) were 0.740-3.400 (mean 1.830) and in control patients (n = 66) 0.090-0.850 (mean 0.320). Using an arbitrary cut-off level of 0.700, the sensitivity of the present autoantibody test was 100%, specificity 91% and positive predictive value 85%. Our identified autoantigens may generate the production of the classical tissue antibodies, known as anti-reticulin and anti-endomysium antibodies, and may be used as antigen in an immunoassay for the antibodies.