Abstract：The primers were designed according to the OMP25 and OMP31 gene sequence from the genome of Brucella suis S2 strain in GenBank, and the 573 bp and 783 bp predicted gene fragments were amplified and cloned respectively into a prokaryotic expression vector pET-28. The recombinant plasmid pET-OMP25 and pET-OMP31 were sucessfully constructed and transformed to bacterium BL21. inducted by IPTG, the recombinant OMP25 and OMP31 protein were expressed mainly in the form of inclusion body. The target proteins were purified and loaded to SDS-PAGE gel for Western blot analysis. The results showed that the purified proteins had good immunological activity. Coating with the two recombinant proteins, the conditions of an indirect ELISA tests were optimized as follows: the best ratio of OMP31 and OMP25 proteins is 4:1, the antigen concentration is 10μg/mL, the best dilution of serum is 1:50, and the critical value is 0.314. based on the conditions above, the sheep brucellosis antibody indirect ELISA diagnostic method was established, the diagnostic kit was assembled and further tested for specificity, sensitivity, repeatability, coincidence rate, cross reactivity, etc. In conclusion, the sheep brucellosis antibody ELISA diagnostic kit has good specificity,sensitivity, and repeatability, and it can reach the quality of similar commercial kits. Therefore, it can be applied in field detection.
王海丽, 董炳梅, 李芬, 许崇友, 王金良. 布鲁氏菌OMP25与OMP31蛋白的表达及其间接ELISA诊断试剂盒研制[J]. 中国人兽共患病学报, 2019, 35(5): 404-410.
WANG Hai-li, DONG Bing-mei, LI Fen, XU Chong-you, WANG Jin-liang. Expression of OMP25 and OMP31 proteins of Brucella suis and development of indirect ELISA antibody detection kit. Chinese Journal of Zoonoses, 2019, 35(5): 404-410.
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2019, Vol. 35 
