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目的对原核系统表达的重组刚地弓形虫Peroxiredoxin(rTgPrx)蛋白进行镍亲和层析纯化,免疫大鼠,制备抗血清。方法优化rTgPrx表达条件,获得大量的可溶性蛋白,经镍亲和层析法纯化后皮下注射免疫Wistar大鼠,间接ELISA法测定血清抗体效价。结果在表达菌液A600为0.5,IPTG浓度为0.2 mmol/L,37℃诱导10 h时,可溶性rTgPrx表达量较高;用镍亲和层析纯化获得纯的目的蛋白,免疫大鼠后诱导产生高滴度抗体血清,效价达1∶12 800以上。结论镍亲和层析法纯化的rTgPrx具有免疫原性,用该蛋白免疫大鼠可获得高效价的抗rTgPrx血清。
Objective To purify the recombinant Toxoplasma gondii Peroxiredoxin (rTgPrx) protein expressed in prokaryotic system by nickel affinity chromatography and immunize rats to prepare antiserum. Methods The expression of rTgPrx was optimized and a large number of soluble proteins were obtained. After purified by nickel affinity chromatography, the Wistar rats were injected subcutaneously and the antibody titer was measured by indirect ELISA. Results The expression level of soluble rTgPrx was higher when the A600 concentration was 0.5, the IPTG concentration was 0.2 mmol / L, and the induction time was 10 h at 37 ℃. Pure recombinant protein was purified by nickel affinity chromatography and then induced by immunofluorescence High titer antibody serum, titer up to 1:12 800 above. Conclusion The rTgPrx purified by nickel affinity chromatography is immunogenic, and high titer anti-rTgPrx serum can be obtained by immunization of rat with this protein.