Study On The IELISA Detection Method Of Immune Antibody Against Brucella S2 Attenuated Vaccine

Brucellosis is a chronic zoonotic infectious disease caused by Brucella app,which is a global epidemic.The disease not only brings huge economic losses to the animal husbandry,but also poses a threat to human life.Existing diagnostic methods of brucellosis cannot identify natural infections and vaccine immunity,however,the relevant studies are of great significance.This study screened out the unique,immunogenic three candidate antigen genes from the whole genome sequence of Brucella S2 vaccine and performed bioinformatics analysis,recombinant expression and immunogenicity identification,using the recombinant proteins S2RP2and S2RP3 as antigens and established a differential diagnosis method for Brucella infectivity and S2 vaccine immunological infection.The main results are as follows:(1)The encoded products of S2JY1,S2JY2,S2JY3 genes of Brucella S2vaccine were all non-transmembrane,hydrophilic proteins without hydrophilic peptides;S2JY1,S2JY3 encoded products are stable proteins,S2JY2 encoded products are unstable proteins;The coding proteins of the three genes have 7,19and15antigenic determinants respectively.(2)After the detection of SDS gel electrophoresis,the recombinant expression bacteria BL21(S2JY2)and BL21(S2JY3)were mainly expressed in the form of inclusion body,and the expression level of the recombinant expression strain BL21(S2JY1)was low;The recombinant proteins S2RP2 and S2RP3 were purified with Ni~+sepharose,and higher purity recombinant proteins were obtained(3)The Western-Blot assay showed that the recombinant proteins S2RP2 and S2RP3both reacted with the S2 vaccine immune serum and did not react with sheep naturally infected serum.It had high specificity and immunogenicity.(4)Using recombinant proteins S2RP2 and S2RP3 as antigens,an iELISA differential diagnosis method for Brucella was established.The sensitivity and specificity of the recombinant antigens S2RP2 and S2RP3 were all above 95%.There was no significant difference between the positive detectable rates of the two detection antigens(P?0..05),indicating that the recombinant antigens S2RP2 and S2RP3 can be used to detect antibodies against Brucella S2 vaccine.