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目的:表达人促血液血管细胞生成素(hemangiopoi-etin,HAPO)蛋白并制备兔抗HAPO抗体。方法:利用硫氧还原蛋白TrxA融合表达载体(pET32c),表达可溶性HAPO。表达的HAPO融合蛋白经亲和层析快速纯化,并通过Westernblot进行鉴定。以纯化的人HAPO免疫新西兰大白兔,制备抗体并测定其效价。结果:获得高效表达并纯化的HAPO蛋白。制备的兔抗人HAPO抗体的免疫双扩散的效价为1∶8。结论:建立了HAPO基因的原核表达载体和快速纯化体系并制备出兔抗HAPO的抗体,为研制检测HAPO的ELISA试剂盒奠定了基础。
Objective: To express human hemangiopoi-etin (HAPO) protein and prepare rabbit anti-HAPO antibody. Methods: Soluble HAPO was expressed by using TrxA fusion expression vector (pET32c). The expressed HAPO fusion protein was rapidly purified by affinity chromatography and identified by Western blot. New Zealand white rabbits were immunized with purified human HAPO to prepare antibodies and determine their potency. Results: Highly expressed and purified HAPO protein was obtained. The prepared rabbit anti-human HAPO antibody double immunodouble titer was 1: 8. CONCLUSION: The prokaryotic expression vector and rapid purification system of HAPO gene were established and the rabbit anti-HAPO antibody was prepared, which laid the foundation for the development of an ELISA kit for detecting HAPO.