Study On Intestinal Anaphylaxis Induced By Soybean Protein In Piglet And Detection Technique Of The Allergen

The purpose of this paper was to study effects of anaphylaxis induced by soybean protein on growth performance and intestinal function of piglet, analyze the main allergic components of soybean protein, identify the IgE epitopes, and prepare the McAbs against soybean β-conglycinin. There are three experiments in this paper.In the first experiment,8litters of piglets from sows with same breed and similar parity were randomly divided into two groups, four litters each. One group as the control was fed the diet without soybean; the other as the treatment group was fed the diet containing32%dehulled soybean meal. Trial lasted from7to49day of age, and piglets were weaned at28day of age. The results showed that, comparing with the piglets in control group,(1) the average daily weight gain in treatment group was reduced by37.5%(P<0.01), the feed/gain ratio increased by59.9%(P<0.05).(2) at28and35day of age, serum diamine oxidase activity of treatment group was increased by22.9%(P<0.05) and13.6%(P<0.05) respectively, serum endotoxin concentration was increased by37.0%(P<0.05) and20.0%(P>0.05), and serum D-lactic acid concentration was increased by30.0%(P<0.05) and28.0%(P<0.05); while serum D-Xylose concentration was reduced by29.0%(P<0.05) and11.1%(P>0.05). These results suggested that soybean protein hypersensitivity damaged piglet’s intestinal function, reduced the absorption capacity of nutrients, resulted in harmful substances entering into the body and ultimately decreased growth performance of the piglets.In the second experiment, the allergic serum samples from piglets were obtained by an indirect ELISA method, with which allergenicity of soybean protein was analyzed by Western blot. Seven small peptides were synthesized according to the amino acid sequence ofasubunit of β-conglycinin, then the peptides which contains IgE epitopes were identified by indirect ELISA. The results of western blot showed that β-conglycinin was the major allergenic component of soybean protein, three subunits of which(α、α、β) could react with the allergic serum. The results of indirect ELISA showed that seven peptides could react with the IgG components of the allergic serum, the highest OD value appeared in peptide7; four peptides could react with the IgE components of the allergic serum, and the OD value of peptide7was also the highest. From the above, the peptide7was a desired antigen for preparation of monoclonal antibody.In the third experiment, the BALB/c mice were immunized with a small peptide with IgE epitope obtained from the second experiment. Then spleen cells from immunized mice were fused with SP2/0myeloma cells, and the positive clones which secreted McAbs against were screened by ELISA. The results of western blot showed that the McAb could bind to native β-conglycinin specifically. The result of indirect ELISA showed that the titer of ascites was1:32000by ELISA. The immunoglobulin subtype of the McAb was IgG1, and its light chain was κ-chain. The successful preparation of anti-β-conglycinin McAb has made foundation for establishment of method to detect the allergen content in soybean products based on IgE epitope.