食物致敏性评价表达人IgE高亲和力受体αβγ的大鼠嗜碱性白血病粒细胞2H3细胞株的建立及鉴定

Construction and identification of stable hFcεRIαβγ/RBL-2H3 cells for food allergy assessment

目的建立稳定表达人IgE高亲和力受体alpha、beta和gamma链(human high affinity receptor containing alpha, beta and gamma chain,hFcεRIαβγ)的大鼠嗜碱性白血病粒细胞(rat basophil leukemia,RBL)-2H3细胞株。方法将包装好的载有hFcεRIαβγ慢病毒感染目的细胞RBL-2H3细胞,实时PCR检测细胞hFcεRIαβγ的表达水平,流式细胞仪分选hFcεRIα表达阳性细胞,并检测分选后细胞hFcεRIα的表达水平。结果成功构建重组慢病毒表达载体GV348-hFcεRIαβγ和慢病毒颗粒;慢病毒能够有效感染RBL-2H3细胞,hFcεRIαβγ在RBL-2H3细胞中成功表达,hFcεRIα在RBL-2H3细胞蛋白水平成功表达。结论初步构建了hFcεRIαβγ/RBL-2H3细胞,相较于RBL-2H3细胞,该细胞表达hFcεRIα,可与人血清IgE特异性结合,能够更好地应用于食品致敏性评价体系中。

OBJECTIVE To establish an rat basophil leukemia(RBL)-2H3 cell line stably expressing human high affinity receptor containing alpha, beta and gamma chain(hFcεRIαβγ), in order to provide experimental materials for evaluating allergenicity of food. METHODS The lentivirus was transfected into RBL-2H3 cells, and the mRNA expression of hFcεRIαβγ in cells was detected by real-time PCR and the protein expression of hFcεRIα was detected by flow cytometry. RESULTS Sequencing result showed that recombinant lentiviral vector GV367-hFcεRIαβγ was successfully constructed. According to the result of experiments, lentivirus could effectively infect RBL-2H3 cells. The mRNA of hFcεRIαβγ and protein levels of hFcεRIα in RBL-2H3 cells were successfully overexpressed. CONCLUSION The hFcεRIαβγ/RBL-2H3 cells were preliminarily constructed, which could be binded with human IgE and further used in the evaluation system of food allergy,compared to RBL-2H3 cells.