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人睫状神经营养因子（ｈＣＮＴＦ）克隆入ｐＢＶ２２０中，在ＤＨ５α菌株中表达，重组蛋白以包含体的形式存在，表达量为菌体总蛋白的５０％左右。经比较发现用２ｍｏｌ／Ｌ脲洗涤包含体可溶解大量可溶性细菌蛋白，且包含体损失较小。在高浓度变性剂条件下进行ｓｅｐｈａｒｃｙｌＳ－２００凝胶过滤，解决了纯化中ｈＣＮＴＦ易聚合的问题，在低浓度变性剂条件下进行ＤＥＡＥ离子交换，有利于蛋白活性的保持。经两步纯化后得到均一性ｈＣＮＴＦ，纯度达９５％以上。在自然状态下使ｈＣＮＴＦ复性。纯化复性后的ｈＣＮＴＦ对无血清培养的鸡胚背根节神经元和脊髓腹角运动神经元有明显的维持存活和促进生长发育的生物效应。
Human ciliary neurotrophic factor (hCNTF) was cloned into pBV220 and expressed in DH5α strain. The recombinant protein existed in the form of inclusion body, and the expression level was about 50% of the total bacterial proteins. The inclusion body was found to be soluble in a large amount of soluble bacterial protein with 2 mol / L urea, with less loss of inclusion body. Sepharcyl S-200 gel filtration under high concentration of denaturing agent solves the problem of easy polymerisation of hCNTF during purification, and DEAE ion exchange under low concentration of denaturant is favorable for the maintenance of protein activity. Homogeneous hCNTF was obtained after two steps of purification, with a purity of more than 95%. HCNTF refolding in its natural state. Purified and renatured hCNTF could significantly inhibit the survival of cultured dorsal root ganglion neurons and spinal cord ventral horn motor neurons in serum-free medium.