Analysis of anti-prolamin monoclonal antibody reactivity using prolamin fractions purified by preparative electrophoresis

Coeliac disease (CD) is a gastrointestinal afliction triggered by the inges tion of prolamins from wheat, barley, rye and possibly oats. The only treat ment for CD is a strict diet, free of the toxic components. Immunochemical methods are usually applied to certify foods aimed at coeliac patients. The characterization of a panel of four anti-prolamin monoclonal antibodies (M Abs) to be used to certify gluten-free products is described here. To this aim, purified gliadin, secalin and hordein fractions were obtained by prepa rative electrophoresis at acid pH. This procedure provides purified fractio ns not exposed to denaturing conditions. The specificity of the MAbs was te sted by ELISA against purified fractions and ethanol extracts of wheat, bar ley, rye, oats, rice, maize, buckwheat, sorghum and soy. The four MAbs reco gnized only coeliac-toxic cereals. Each MAb reacted strongly wit gliadins a nd showed differential reactivity against the different prolamin purified f ractions. Some MAbs showed a broad pattern of recognition whereas others pr esented a more restricted one. The reactivity observed corresponded to stru ctural homologies among gliadin, secalin and hordein fractions. It is remar kable that some fractions obtained by electrophoresis in the presence of so dium dodecylsulphate were not recognized by some MAbs, whereas the same com ponents obtained by preparative A-PAGE showed high reactivity. This reinfor ces the suitability of the purification method employed in this study to is olate prolamin fractions. Using these purified prolamins, characterization of anti-prolamin MAb reactivity was achieved.