The Separation And Purification Of SEB And The Making Of SEB Monoclonal Antibody

Enteropathogenic microorganism in food is the main factor of food poisoning disease, and its symptom is mainly diarrhea, vomit. It is harmful to human's health. Staphylococcus aureus is one of the pathogens for human being and some animals and it can bring food poisoning. Its pathogenicity rest with the ability to secrete pathotoxin and zymins, and heat-resistant Staphylococcus aureus enterotoxin B is the main causation that results in food poisoning and Staphylococcus gastritis and enteritis.Fast and automatic test ways is on the quick developing period. At present, there are many test ways in pathogenic microbiology test and some new quick test ways appear, but, at present, only biology chip can meet well human being's quick, veracious and simple demand for testing pathogenic microbiology. Moreover, gene doesn't correspond to its expressive toxin protein; gene chip's analytic result can not reflect protein's level. So, one of the methods is to study gene's expressive substance—protein.This thesis is to make SEB monoclonal antibody that is used by protein chip and do some anterior work for protein chip's application in pathogenic microbiology. The main process includes the separation and purification of SEB and the making of SEB monoclonal antibody. As to producing toxin S. aureus standard bacterium CMCC (B) 26002, enterotoxin cultivation method use velum cultivation that agar culture dish are covered with dialytic cellophane paper. Enterotoxin is pretreated. The following work is to use positive ion-exchange carboxy methyl cellulose (CMC) to separate SEB, and then use Sephadex G-75 to purify SEB according to filter theory. Thus, we can gain purified SEB only by two steps. In the experiment, Purified SEB was used to immunize BALB/C mouse, then, let spleen cell and myeloma cells (Sp2/0) fuse. After selective cultivation, test and identification, The experiment gain successfully myeloma cells that can secrete SEB monoclonal antibody and its idiosyncrasy is good through enzyme linked immunosorbent assay (ELISA). The results of this experiment have established good foundation for protein chips in testing pathogenic microbiology.