Measuring of IgG2c isotype instead of IgG2a in immunized C57BL/6 mice with Plasmodium vivax TRAP as a subunit vaccine candidate in order to correct interpretation of Th1 versus Th2 immune response
Measuring of IgG2c isotype instead of IgG2a in immunized C57BL/6 mice with Plasmodium vivax TRAP as a subunit vaccine candidate in order to correct interpretation of Th1 versus Th2 immune response
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DOI：
10.1016/j.exppara.2020.107944
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发表时间：
2020-09-01
影响因子：
2.1
通讯作者：
Djadid, Navid Dinparast
中科院分区：
文献类型：
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作者：
Nazeri, Saeed;Zakeri, Sedigheh;Djadid, Navid Dinparast
鼠同种型抗体的评价在亚单位疫苗开发中是必不可少的，因为具有不同遗传背景的近交系小鼠品系对重组蛋白的反应不同。在这方面，本研究的主要目标是测量和比较C57 BL/6小鼠中IgG同种型应答的概况。为此目的，间日疟原虫血小板反应蛋白相关粘附蛋白（PvTRAP）基因的胞外区在大肠杆菌Rosetta（DE 3）-pET 23 a中表达。然后，将重组PvTRAP单独或与弗氏完全佐剂乳化用于免疫C57 BL/6小鼠。评价了重组PvTRAP诱导的抗体和细胞免疫应答的作用。结果显示，在单独接受rPvTRAP的组中，抗rPvTRAP IgG 2c的水平显著高于IgG 2a（平均OD 490分别= 0.798 +/- 0.12和0.39 +/- 0.1）并用CFA/IFA乳化（平均OD 490分别= 1.48 +/-0.07和0.605 +/-0.13; P < 0.05，独立样品t检验）。此外，用rPvTRAP和rPvTRAP + CFA/IFA免疫的小鼠具有中等亲合力IgG 2a抗体，但具有高亲合力IgG 2c抗体，血清抗体滴度的平均值结果显示，在rPvTRAP和rPvTRAP + CFA/IFA小鼠组中，IgG 2a终点滴度（分别为1：3200和1：25，600）显著低于IgG 2c（分别为1：25，600和1：102，400）。此外，结果显示，与小鼠对照组相比，在接受单独的rPvTRAP和与CFA/IFA乳化的rPvTRAP的小鼠组中，诱导显著水平的IFN-γ（P < 0.05，独立样本t检验）和不可检测水平的IL-4。总之，我们的结果表明，为了正确解释C57 BL/6小鼠品系中的Th 1免疫应答，关键是测量IgG 2c而不是IgG 2a沿着IFN-γ。
Evaluation of the murine isotype antibodies is essential in subunit vaccine development because inbred mouse strains with diverse genetic backgrounds respond different to recombinant proteins. In this regard, the main goal of this study was to measuring and comparing the profile of IgG isotype responses in C57BL/6 mice. For this purpose, the extracellular region of plasmodium vivax thrombospondin-related adhesive protein (PvTRAP) gene was expressed in Escherichia coli Rosetta (DE3)-pET23a. Then, the recombinant PvTRAP alone or emulsified with Freund's complete adjuvant were applied for immunization of the C57BL/6 mice. The role of antibodies and cellular immune responses induced by recombinant PvTRAP were evaluated. The results showed the level of anti-rPvTRAP IgG2c was significantly higher than IgG2a in the groups that received rPvTRAP alone (mean OD490 = 0.798 +/- 0.12 and 0.39 +/- 0.1, respectively) and emulsified with CFA/IFA (mean OD490 = 1.48 +/- 0.07 and 0.605 +/- 0.13, respectively; P < 0.05, independent sample t-test). Additionally, the immunized mice with rPvTRAP and rPvTRAP + CFA/IFA had an intermediate-avidity IgG2a antibody but high-avidity IgG2c antibody as well as the mean of serum antibody titers results exhibited that in both rPvTRAP and rPvTRAP + CFA/IFA mouse groups, IgG2a end-point titer (1:3200 and 1:25,600, respectively) was noteworthy lower than IgG2c (1:25,600 and 1:102,400, respectively). Moreover, the results revealed the eliciting significant levels of IFN-gamma (P < 0.05, independent sample t-test) and no detectable level of IL-4 in the mouse groups received rPvTRAP alone and emulsified with CFA/IFA as compared to the mouse control groups. In general, our results showed that for correctly interpreting of Th1 immune responses in C57BL/6 mouse strain it is critical to measure IgG2c instead of IgG2a along with IFN-gamma.