本文選題：貓杯狀病毒　切入點(diǎn)：VP-E蛋白　出處：《中國預(yù)防獸醫(yī)學(xué)報》2017年12期

【摘要】：為制備抗貓杯狀病毒(FCV)衣殼蛋白VP1的單克隆抗體(MAb)及鑒定其表位,本研究以原核表達(dá)的重組VP1-E蛋白(aa427~aa524)免疫BALB/c小鼠,取其脾細(xì)胞與骨髓瘤細(xì)胞(SP2/0)進(jìn)行融合制備雜交瘤細(xì)胞;以純化的FCV-2280病毒粒子作為檢測抗原,采用間接ELISA檢測方法篩選雜交瘤細(xì)胞株,分析雜交瘤細(xì)胞所分泌的抗體亞型、中和活性及所識別的抗原表位等。結(jié)果表明,獲得兩株穩(wěn)定分泌抗FCV VP1-E MAbs的雜交瘤細(xì)胞E2F1和E2F6,抗體均為IgG1亞型,輕鏈為κ鏈。雜交瘤細(xì)胞分泌的抗體無中和活性。Western blot結(jié)果顯示,兩種抗體可以特異性識別FCV-2280,表明其針對的抗原表位為線性表位。兩株MAb識別VP1-E序列均為~(467)YICGSLQRAWG~(477)。生物信息學(xué)分析顯示該抗原表位在FCV VP1蛋白中相對保守。該MAb為建立特異性強(qiáng)、靈敏度高的FCV的檢測方法奠定基礎(chǔ)。
[Abstract]:In order to prepare monoclonal antibody against capsid protein VP1 and identify its epitopes, BALB/c mice were immunized with prokaryotic expressed recombinant VP1-E protein (aa427a524). The spleen cells were fused with myeloma cells (SP2 / 0) to produce hybridoma cells.Using purified FCV-2280 virus particles as detection antigen, indirect ELISA assay was used to screen hybridoma cell lines, and antibody subtypes, neutralizing activities and antigen epitopes recognized by hybridoma cells were analyzed.The results showed that two hybridoma cells, E2F1 and E2F6 secreted stably against FCV VP1-E MAbs, were obtained. The antibody was IgG1 subtype and the light chain was 魏 chain.The antibody secreted by hybridoma cells had no neutralization activity. Western blot results showed that the two antibodies could specifically recognize FCV-2280, indicating that the antigenic epitopes were linear epitopes.The VP1-E sequences of the two strains identified by MAb were both 467 (YICGSLQRAWGN).Bioinformatics analysis showed that the epitope was relatively conserved in FCV VP1 protein.The MAb lays a foundation for the establishment of a specific and sensitive method for the detection of FCV.
【作者單位】： 中國農(nóng)業(yè)科學(xué)院哈爾濱獸醫(yī)研究所獸醫(yī)生物技術(shù)國家重點(diǎn)實(shí)驗(yàn)室;
【基金】：國家重點(diǎn)研發(fā)項目(2016YFD0501001) 中央級公益性科研院所基本科研業(yè)務(wù)費(fèi)專項(1610302017012)
【分類號】：S852.65

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