Food Allergen Labeling Regulations in Japan require mandatory labeling of seven allergens in foods at 10 µg/g or more, including buckwheat proteins. Allergenic contamination of food is usually monitored using enzyme-linked-immunosorbent-assay (ELISA), which requires multiple handling steps, an enzyme-labeled secondary antibody, and optical equipment for quantification. In contrast, a FET sensor and an immunochromatography assay strip are potential assay tools without those drawbacks due to their direct sensing mechanism.
In this study, we propose a FET sensor and an immunochromatography assay strip be devised for detection of buckwheat protein. Here we attempt to improve the sensitivity of a FET sensor for buckwheat protein BWp16 by increasing the net charge of its molecules.
BWp16 was determined specifically using a FET sensor. The response of the FET sensor was improved markedly by pretreatment of recombinant BWp16 (rBWp16) with SDS. The response of the FET sensor was correlated positively with the concentration of rBWp16 down to a detection limit below1 ng/ml.
The charge number of BWp16 calculated from its molecular weight (15,106 Da) and isoelectric point (pH 5.25) was small (−3.67/molecule) at the sensing condition of pH 7.4 used with the FET sensor. After the pretreatment with SDS, BWp16 was covered with enough negative charge to increase sensitivity to approximately 10 ng of buckwheat proteins/g of food material.
We devised an immunochromatography assay strip and tested the strip on 40 food samples. The assay strip gave a lower limit of detection below 5ng/ml of BWp16.
SDS pretreatment of BWp16 protein from buckwheat increased its sensitivity to detection with a FET sensor nearly 1000-fold. This simple charge-amplification technique can increase the sensitivity of a FET sensor to a targeted protein that would ordinarily carry a small net charge.
Both a FET sensor and an immunochromatography assay strip achieved highly sensitive and specific detection of buckwheat protein.