The Preparation And Therapeutic Roles Of Single-chain Antibody That Against Hla Of Stapylococcus Aureus To Control Mastitis In Mice

Bovine mastitis is a frequently occurring disease that affects the development of the dairy industry and leads to huge losses in dairy production.Staphylococcus aureus is the main pathogens.At present,the clinical treatment of mastitis caused by Staphylococcus aureus mostly uses Chinese herbal medicine,inactivated vaccine and antibiotic treatment,but adverse reactions such as "resistant milk" and drug-resistant strains have prompted people to seek better solutions.However,vaccines against S.aureus fail to provide adequate protection for cow mastitis.With the development of immunology and molecular biology,antibody preparations are favored by more researchers,especially single chain antibodies,which not only remain intact.Single chain antibodies(scFv)have the advantages of maintaining intact antigen-antibody binding sites,but with a molecular weight being only 1/6 of intact antibodies,and also have low immunogenicity and strong tissue penetration.Single-chain antibodies(scFv)have been successfully used in antibody therapy and have shown evident effects.In this study,the virulence factor-? hemolysin(Hla),a critical virulence factor affecting its pathogenicity,was used to amplify the Hla encoding gene from Staphylococcus aureus DNA,and then transformed into E.coli for expression and purification.The prokaryotic expression product(rHla),was further employed as antigen,and a high affinity scFv,for rHla was screened from bovine phage scFv expression library.The scFv was expressed in prokaryotic expression and purified scFv protein(scFvZZ107)was obtained.In order to verify the protective effect of ? hemolysin single chain antibody(scFv),in the mastitis model of infected Staphylococcus aureus in lactation stage,different concentrations of scFvZZ107,were observed by bacterial count and pathological sections.The levels of cytokines and apoptosis of breast tissues were measured to verify the protective effect of the scFv on mastitis induced by Staphylococcus aureus in mice.Compared with the negative control group,the scfv group with different dose pairs had a protective effect on intramammary infection,and the number of bacteria in the medium and high dose scFv test group was significantly reduced(p < 0.05),and the breast tissue was relatively intact.The contents of IL-1?,IL-1?,IL-8 and IL-10 decreased significantly(p < 0.05),indicating that ?-hemolysin scfv may have protective effect on mastitis in mice.On the one hand,the results of this study provide a potential high-quality antibody molecule for the treatment of cow mastitis induced by Staphylococcus aureus,on the other hand,provide a theoretical basis for studying the pathogenesis of ? hemolysin at the molecular level.