發(fā)布時間：2018-05-01 22:59

  本文選題：IL-a + IL-p多克隆抗體　； 參考：《南昌大學學報(理科版)》2017年01期

【摘要】：利用分子克隆方法構建重組載體pGEX-4T-1-IL-12a,轉化BL21(DE3),經IPTG誘導表達,谷胱甘肽瓊脂糖凝膠4B親和層析柱純化后,獲得重組融合蛋白GST-IL-12p35。以此蛋白為免疫抗原免疫新西蘭大耳白兔制備IL-12p35多克隆抗體。采用ELISA法評估抗體效價,Western blot檢測抗體特異性。LPS刺激小鼠脾細胞,ELISA法檢測IL-12抗體阻斷后細胞培養(yǎng)上清中的IFN-γ的分泌量。結果:成功構建重組載體pGEX-4T-1-IL-12a,并獲得重組融合蛋白GST-IL-12p35(約48kDa)�？乖庖咝挛魈m大耳白兔制備IL-12p35多克隆抗體,ELISA法測得IL-12p35多克隆抗體效價為1鐩256 000,Western blot證實制備的多克隆抗體能夠識別結合重組蛋白。IL-12p35多克隆抗體能夠有效地抑制LPS誘導的脾細胞IFN-γ的表達。
[Abstract]:The recombinant plasmid pGEX-4T-1-IL-12awas constructed by molecular cloning and transformed into BL21DE-3. The recombinant fusion protein GST-IL-12p35 was obtained by IPTG induced expression and purified by glutathione agarose gel 4B affinity chromatography. IL-12p35 polyclonal antibodies were prepared by immunizing New Zealand rabbits with this protein. ELISA assay was used to evaluate the titer of antibody. Western blot was used to detect the secretion of IFN- 緯 in the culture supernatant of murine spleen cells stimulated by IL-12 antibody. Results: the recombinant vector pGEX-4T-1-IL-12awas successfully constructed and the recombinant fusion protein GST-IL-12p35 was obtained. Antigen-immunized New Zealand white rabbits with IL-12p35 polyclonal antibodies were obtained by Elisa. The titer of IL-12p35 polyclonal antibodies was 1: 256 000. Western blot confirmed that the prepared polyclonal antibodies could recognize the binding protein. IL-12p35 polyclonal antibodies could be effectively inhibited. LPS induced expression of IFN- 緯 in splenocytes.
【作者單位】： 鄭州大學生命科學學院分子免疫實驗室;鄭州人民醫(yī)院;河南省醫(yī)藥科學研究院;
【基金】：重大新藥創(chuàng)制科技重大專項(2012ZX09103301-022) 國家自然科學基金資助項目(U1204817、81373119、81571526)
【分類號】：S852.4

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