Enzyme Linked Immunosorbent Assay Staphylococcal Enterotoxin B From Milk

Enzyme-linked immμnosorbent assay (ELISA) was developed for analysing Staphylococcal enterotoxin B (SEB) from milk using the anti-SEB antibody. Main research contents were followed:1. pμrification for SEBA method was established to purify Staphylococcal enterotoxin B from the broth of Staphylococcus aureus BP1. SEB was separated and purified successively by ultrafiltration, salting out with ammonium sulfate, ionexchange chromatography and molecular exclusion chromatography. The total yield was 20% with the purity of 90%.2 Production of anti-SEB antibodyBALB/c mice were injected with the SEB. The antibody began to obviously produce in mouse when the coarse antigen was injected after 90 days, while the antibody began to obviously produce in mouse when the pure antigen was injected after 60 days, when the antibody reached to peak, then began to go down. The highest titer of antibody at peak was1:100,000 using the indirect competitive ELISA.3 ELISA detection of SEB from milkThe indirect competitive ELISA (ICELISA ) was used to analyze the SEB content in milk. The results showed that the detecting range was 1~1000ng/ml, and the recovery of this method was 94.5%.4 Preparation for ELISA kitsELISA kits were stored at 4℃for two months and -20℃for three months after appropriate pre-treatment, the OD490nm values of kits were detected by indirect ELISA, the results revealed that the OD490nm valμes of kits were 1/10 and 1/3 times of initial OD490nm value, respectively.