抗小鼠IgG纳米抗体的制备及应用

Preparation and application of anti-mouse IgG nanobody

目的构建抗小鼠免疫球蛋白G(immunoglobulin G,IgG)纳米抗体文库,筛选抗小鼠IgG的纳米抗体,并制备小鼠IgG的特异性亲和层析介质。方法以小鼠IgG为靶点,构建纳米抗体文库,利用噬菌体筛选技术对纳米抗体文库进行筛选。得到的纳米抗体经重组表达和纯化后制备成小鼠IgG亲和层析介质,并利用小鼠血清进行测定和验证。结果构建了多样性好且库容量达2.3×107 cfu的初始文库,经3轮淘选后,共筛选出4条有结合活性的纳米抗体序列,其中纳米抗体2E2经预活化填料偶联后,对小鼠IgG载量可达18.55 mg/mL,可用于小鼠IgG的纯化和检测。结论利用噬菌体筛选技术成功筛选出抗小鼠IgG的纳米抗体,并制备出特异性亲和小鼠IgG的亲和层析介质。

Objective To construct the anti-mouse IgG nanobody library,screen anti-mouse IgG nanobody,and prepare the specific affinity chromatography medium of mouse IgG.Methods Nanobody library was constructed using mouse IgG as target and screened by phage library screening technique.The obtained nanobody was subjected to recombinant expression and purification,and prepared into affinity chromatography medium of mouse IgG which was determined and validated by using mouse serum.Results A well-diversified initial library with a capacity of 2.3×107 cfu was constructed.After three rounds of panning,four nanobody sequences with binding activity were obtained,among which nanobody 2E2 after being coupled to the pre-activated medium,with a load of 18.55 mg/mL,could be used for purification and determination of mouse IgG.Conclusion Anti-mouse IgG nanobodies were successfully screened by phage library screening technique,and the affinity chromatography medium with specific affinity to mouse IgG was prepared.