Inter-laboratory optimization of protein extraction, separation, and fluorescent detection of endogenous rice allergens
2016
Satoh, Rie | Teshima, Reiko | Kitta, Kazumi | Lang, Gang-Hua | Schegg, Kathleen | Blumenthal, Kenneth | Hicks, Leslie | Labory-Carcenac, Bénédicte | Rouquié, David | Herman, Rod A. | Herouet-Guicheney, Corinne | Ladics, Gregory S. | McClain, Scott | Poulsen, Lars K. | Privalle, Laura | Ward, Jason M. | Doerrer, Nancy | Rascle, Jean-Baptiste
In rice, several allergens have been identified such as the non-specific lipid transfer protein-1, the α-amylase/trypsin-inhibitors, the α-globulin, the 33 kDa glyoxalase I (Gly I), the 52–63 kDa globulin, and the granule-bound starch synthetase. The goal of the present study was to define optimal rice extraction and detection methods that would allow a sensitive and reproducible measure of several classes of known rice allergens. In a three-laboratory ring-trial experiment, several protein extraction methods were first compared and analyzed by 1D multiplexed SDS-PAGE. In a second phase, an inter-laboratory validation of 2D-DIGE analysis was conducted in five independent laboratories, focusing on three rice allergens (52 kDa globulin, 33 kDa glyoxalase I, and 14–16 kDa α-amylase/trypsin inhibitor family members). The results of the present study indicate that a combination of 1D multiplexed SDS-PAGE and 2D-DIGE methods would be recommended to quantify the various rice allergens.
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