Determination of Anti-?-Gliadin Antibodies in Serologic Tests for Coeliac Disease

BACKGROUND: Serologic detection of coeliac disease in the general population or in subjects belonging to risk groups implies the use of a test with high efficiency, large-scale use, and low cost. The enzyme-linked immunosorbent assay (ELISA) technique is the most appropriate assay for performing this kind of studies. Even though anti-gliadin determination has been the test most frequently used as the first step in screening procedures, many false-positive results produced a low-specificity test. In a previous work a selective recognition of omega-gliadins, mainly by IgA antibodies, was observed. Results also indicated that omega-gliadins can be useful as antigens in serologic detection of coeliac disease. We therefore wanted to analyse the anti-gliadin antibody reactivity by using purified gliadins and to evaluate the actual performance of the anti-omega-gliadin antibody test. METHODS: A population consisting of 105 coeliac patients, 81 healthy controls, and 73 subjects in a disease control group was analysed. Anti-endomysium (EMA), both IgG and IgA anti-omega-gliadins, and anti-tissue transglutaminase (tTG) antibodies were determined. RESULTS: Concordant results, positive and negative, in the EMA and IgG and IgA anti-gliadin determinations were observed in 220 of 259 samples from the total population analysed. The three assays showed high efficiency, being 96.9%, 90.7%, and 91.1% for EMA and anti-omega-gliadins IgG and IgA, respectively. Anti-tTG determination was performed on 103 samples (69 controls and 34 coeliac patients), finding 4 false results (2 false positive and 2 false negative), whereas anti-omega-gliadins showed 10 false results (5 false negative and 5 false positive), 3 of which were coincident with anti-tTG determination. To compare the reactivity of anti-gliadin antibodies, alpha-, beta-, gamma- and omega-gliadins were isolated under non-denaturing conditions by acid preparative electrophoresis and cation-exchange fast protein liquid chromatography (FPLC) and used in an indirect ELISA test. The composition of these fractions was analysed by means of capillary electrophoresis, showing no cross-contamination among them. CONCLUSIONS: The comparison of results using purified gliadins shows that omega-gliadins present a differential reactivity that has not previously been documented. Results using omega-gliadins isolated by either preparative electrophoresis or FPLC were similar. Tests using the purified omega-gliadin fraction present the best performance when anti-gliadin antibodies are evaluated.