抗白念珠菌人鼠嵌合抗体真核表达载体的构建与表达

Construction and Expression of A Human-mouse Chimeric Antibody Against Candida albicans

目的构建抗白念珠菌人鼠嵌合抗体的真核表达载体,并实现真核表达。方法从含有单克隆天然抗白念珠菌抗体3B4可变区基因的载体pMDT-V2H和pUC-VL中PCR克隆单抗3B4的可变区VH和VL基因,依次插入含有人免疫球蛋白κ轻链恒定区基因和γ1重链恒定区基因的真核表达载体pMH-CA。经DNA序列测定正确后,电穿孔转染J558L细胞进行嵌合抗体表达,RT-PCR,ELISA方法对抗体表达进行鉴定。结果成功构建了抗白念珠菌人鼠嵌合抗体,转染真核细胞后RT-PCR显示人鼠嵌合抗体重链和轻链mRNA的转录,ELISA证实了抗白念珠菌人鼠嵌合抗体的表达以及对白念珠菌的识别。结论成功构建抗白念珠菌人鼠嵌合抗体的真核表达载体,并表达出具有结合活性的基因工程抗体。

Objective To construct eukaryotic expression vector for a human-mouse chimeric antibody against C. albicans, and express the antibody in eukaryotic cells. Methods Variable region genes of a mouse hybridoma cell line 3B4 against C. albicans were cloned from plasmid pMDT-V2H and pUC-VL separately, and were inserted into the chimeric antibody eukaryotic expression vector pMH-CA. The constructed vector was transfected into J558L cells, and expression of chimeric antibody was detected by RT-PCR and ELISA. Results Eukaryotic expression vector of the chimeric antibody against C. abicans was successfully constructed. The expression of chimeric antibody was firstly proved by RT-PCR, and the specificity of the chimeric antibody was confirmed by ELISA, which showed that chimeric antibody could bind to both keratin and C. albicans. Conclusion A human-mouse chimeric antibody against C. albicans was constructed successfully. The chimeric antibody has the binding specificity to both self-antigen keratin and C. albicans.