Establishment of a Sandwich ELISA for Detecting Soybean Agglutinin as an Anti-nutritional Factor
To establish a sensitive and specific enzyme-linked immunosorbent assay(ELISA)for the detection of soybean agglutinin(SBA).Mouse-derived monoclonal antibody was used as the capturing antibody and rabbit-derived polyclonal antibody as the detecting antibody,with a sheep anti-rabbit en-zymic conjugate antibody as the secondary antibody.This sandwich method for SBA detection was op-timized in terms of antibody concentration and reaction time.This method was applied to detect SBA levels in differenet soybean products,including soybean,soybean meal,extruded soybean and enzyme hydrolyzed soybean meal.The results showed that the detection range of the developed sandwich ELISA was 39.06-1 250 ng/mL,the detection limit was 3.00 ng/mL,the fortified recovery was 77.96％-116.15％,the coefficient of variation was 6.11％-18.34％.There was no cross-reaction with other anti-nutritional factors such as trypsin inhibitors BBI,KTI,glycinin and β-conglycinin in soybean.The concentration of SBA in soybean was higher than that in soybean meal,extruded soy-bean meal and enzyme hydrolyzed soybean meal,respectively(P＜0.05).SBA in soybean meal was higher than in extruded soybean meal and enzyme hydrolyzed soybean meal,respectively(P＜0.05)and in extruded soybean meal was higher than in enzyme hydrolyzed soybean meal(P＜0.05).These findings indicate that the established SBA sandwich ELISA detection method is both sensitive and spe-cific and can be used for monitoring and detecting soybean agglutinin in feed.
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