Gliadin and glutenin contents are known to affect gluten functional properties. ELISA tests, which are rapid and easy to perform, can provide very convenient methods for industrial quality control. The purpose of this study was to develop a competitive ELISA for quantification of gliadins in wheat flour and compare the results with those obtained by nitrogen and reverse-phase high-performance liquid chromatography (RP-HPLC) determination of gliadin content after sequential extraction. Asαβ-gliadin and total gliadin contents are highly correlated, polyclonal antibodies directed against the C-terminal peptide ofαβ-gliadins were used in a sequential competitive ELISA to quantify total gliadins. In 21 flour samples, the results with ELISA were closely correlated with those obtained by nitrogen and RP-HPLC determinations (r> 0.82,P< 0.001), indicating that the ELISA test based on detection ofαβ-gliadins allowed fairly accurate quantification of total gliadins. Immunoblotting analysis after acid-PAGE showed that someβ-gliadin components were poorly detected or undetected by the antiserum, which could account for differences between the immunochemical and biochemical values observed for a few cultivars.