A Rapid Immunomagnetic Bead-based Chemiluminescence For The Separation,Concentration And Detection Of Staphylococcal Enterotoxin B

Staphylococcal enterotoxin?SE?is a series of exotoxins secreted by Staphylococcus aureus.SE is a single peptide chain soluble protein,molecular weight 27-30KDa.SE has four common characteristics:1,cause vomiting and gastroenteritis;2,superantigen;3,heat and pepsin resistance;4,similar tertiary structure.SE is the causative substance basis of Staphylococcal food poisoning?SPF?and participates in the occurrence and development of sepsis-induced multi-organ failure?MOF?.SE has high toxicity,good stability and simple preparation.SEB is listed by the United States as the biological warfare agent most likely to be used for biological terrorist activities.This study established a rapid detection method based on immune magnetic separation and chemiluminescence detection of Staphylococcus aureus enterotoxin?SEB?,including three aspects:Preparation of Staphylococcus aureus Type B enterotoxin:The existing engineering bacteria were cultured in large quantities to separate and purify the broken cells,so as to prepare for the subsequent preparation of the SEB rabbit polyclonal antibody.Staphylococcus aureus B enterotoxin rabbit polyclonal antibody preparation:The purified Staphylococcus aureus B enterotoxin routine immunization of large white rabbits,the serum reaches the titer after the carotid artery blood,the serum was isolated and purified.Establishment of a method for the detection of Staphylococcus aureus enterotoxin B based on immunomagnetic separation and chemiluminescence double-antibody sandwich method:An immunomagnetic beads-based chemiluminescence?IMBs-CLIA?was developed for the detection of Staphylococcal enterotoxin B?SEB?.It is a system for the efficient separation and concentration of SEB using biologically prepared immunomagnetic beads,Immunomagnetic beads?IMBs?were employed as the solid phase.The anti-SEB monoclonal antibody?Mc Ab?bound to IMBs was used as capture probe and an anti-SEB polyclonal antibody?PcAb?from goat as primary antibody?PA?has a function to link SEB and detector probe.Rabbit anti-goat antibody,labelled with horseradish peroxidase?HRP?as second antibody?SA?,was employed as detector probe and enhance the detection signal.Hence,a sandwich complex is formed as“IMBs–SEB–PA-SA”,which significantly shortened detection compared with classic sandwich ELISA and only need 56 minutes to complete the whole process.Experimental results indicate that this detection platform exhibits good sensitivity for SEB with a detection linear range(0.8–500ng m L^-1).The proposed assay is thus a highly promising alternative method for detecting the contamination of SEB in the food industry.