Preparation and immunogenicity of a polyclonal antibody against outer membrane protein OspC and flagellin FlaB of Borrelia burgdorferi
ZHANG Xue-chao1, ZHU Han-ping2, YAO Ping-ping2, XU Hai-jun3, XU Fang2, SUN Yi-sheng2, LU Hang-jing2, ZHANG Yun4, YUE Ming5, YANG Zhang-nyu2,4
1 Hangzhou Center for Disease Control and Prevention, Hangzhou 310021, Zhejiang Province, China; 2 Zhejiang Provincial Center for Disease Control and Prevention; 3 Institute of Insect Sciences, Zhejiang University; 4 Institute of Military Medicine, Eastern Theater of the Chinese People's Liberation Army; 5 The First Affiliated Hospital of Nanjing Medical University
Abstract：Objective To prepare a polyclonal antibody against the outer membrane protein OspC and the flagellin FlaB of Borrelia burgdorferi (also called Lyme disease spirochete) after prokaryotic expression, and to test its immunogenicity. Methods The genomic DNA of B.garinii strain BgNMJW1 was used as a template to amplify the gene segments of OspC and FlaB using polymerase chain reaction (PCR); then the PCR products were subcloned into the expression vector pGEX-6p-1 and transformed into the expression strain Rosetta. The fusion proteins were expressed after induction with isopropyl thiogalactoside (IPTG) and purified using glutathione transferase (GST) column or via gel cutting. The purified proteins were then used to immunize New Zealand white rabbits to obtain polyclonal antiserum. Results The result of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed that compared with the control, the vector carrying the target genes had obvious expression of recombinant proteins after induction, and the sizes of the recombinant proteins were about 49×103 and 63×103, respectively. The induction result showed that the expression of the induced proteins reached its peak level if 1 mmol/L IPTG was added when the bacteria were cultured to an absorbance (A) of 0.4, followed by inducing at 25℃ for 10 hours. By immunizing New Zealand white rabbits with the purified fusion proteins, the polyclonal antiserum was obtained and used to detect OspC and Flab of B.garinii strain BgNMJW1 and B.burgdorferi strain BbB31A3 in Western blot, then the clear detection bands were obtained. Conclusion The combined application of OspC and FlaB can play an effective role in the diagnosis of Lyme disease.
张学潮, 朱函坪, 姚苹苹, 徐海君, 徐芳, 孙一晟, 卢杭景, 张云, 岳明, 杨章女. 伯氏疏螺旋体外膜蛋白OspC和鞭毛蛋白FlaB多克隆抗体的制备和免疫原性研究[J]. 中国媒介生物学及控制杂志, 2020, 31(5): 531-535.
ZHANG Xue-chao, ZHU Han-ping, YAO Ping-ping, XU Hai-jun, XU Fang, SUN Yi-sheng, LU Hang-jing, ZHANG Yun, YUE Ming, YANG Zhang-nyu. Preparation and immunogenicity of a polyclonal antibody against outer membrane protein OspC and flagellin FlaB of Borrelia burgdorferi. Chines Journal of Vector Biology and Control, 2020, 31(5): 531-535.
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