Capparelli, Rosanna, A., Costabile, M., Viscardi, I., Ventimiglia, L., Longobardi, D., Fenizia, D., Iannelli, Costabile, A, Viscardi, M, Ventimiglia, I, Longobardi, L, Fenizia, D, Iannelli, Domenico
Gliadin is a heterogeneous group of alcohol-soluble wheat storage proteins, comprising ≈50% of the gluten proteins (Campbell et al 1987; Fido et al 1997). Gliadin influences important baking properties of wheat, in particular loaf volume (Weegels et al 1994; Khatkar et al 2002) and water absorption (Dong et al 1992). Gliadin has been used also as a tool for wheat varietal identification (Bietz et al 1984). More importantly, gliadin, along with the prolamins of rye, barley and possibly of spelta wheat (Kasarda and D’Ovidio 1999), is responsible for the pathogenesis of the celiac disease. This food intolerance has a prevalence of 1/200 to 1/300 (Hin et al 1999) and is strongly associated with DQ2 and DQ8 HLA class II haplotypes (Sollid 2000). The symptoms of the disease disappear when patients follow a strict gluten-free diet and reappear when gluten is introduced again. Thus, treatment of celiac disease relies exclusively on a gluten-free diet. At present, ELISA is used frequently to ascertain the absence of toxic prolamins in gluten-free foods (Sorell et al 1998). We investigated the possibility of using ELISA for flow cytometry. The applications of this technique are becoming increasingly numerous. However, flow cytometry can only analyze single cells. To mimic a cell population in this study, the gliadin in test samples was extracted with 70% ethanol, adsorbed on latex particles, coated with rat anti-gliadin antibodies, labeled with fluoresceinated goat antirat immunoglobulins, and finally measured by flow cytometry. The same approach has been used before to detect toxins (Evidente et al 1997), antibodies (Presani et al 1989; Iannelli et al 1998), and viruses (Iannelli et al 1996), which are all not visible to the laser in their natural state. The method described in this study can detect all cereal prolamins (gliadin, secalin, hordein, and avenin) potentially toxic to the celiac patient with a sensitivity of 5 ppm (5 ng of gliadin added to 1 mg of maize flour), introducing the condition to further reduce the amount of gluten allowed in gluten-free foods.
