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目的对单甲氧基聚乙二醇丁醛(mPEG-ButyrALD,相对分子质量20×103)修饰重组人睫状神经因子(rh-CNTF)的条件进行优化。方法优化了pH值、温度、时间、rh-CNTF与PEG的修饰比例、rh-CNTF浓度等修饰条件,并对修饰产物行分离纯化。鸡胚背根节培养法检测和比较rh-CNTF和PEG-rhCNTF的体外生物学活性。结果rh-CNTF与PEG的修饰比例、rh-CNTF的浓度和pH值是主要影响因素,最佳pH为6.5,最佳rh-CNTF浓度为1～2 mg/ml,rh-CNTF与PEG的最佳质量比为1∶1,时间和温度影响甚微。rh-CNTF的体外神经营养活性约为PEG-rhCNTF的2倍。结论本法优化了修饰重组人睫状神经因子的条件。
Objective To optimize the conditions of recombinant human ciliary neurotrophic factor (rh-CNTF) with monomethoxy polyethylene glycol butyral (mPEG-ButyrALD, molecular weight 20 × 103). The optimized conditions of pH value, temperature, time, modification ratio of rh-CNTF and PEG, concentration of rh-CNTF and so on were optimized. The modified products were isolated and purified. The chicken embryo dorsal root ganglia culture method was used to detect and compare the in vitro biological activities of rh-CNTF and PEG-rhCNTF. Results The modification ratio of rh-CNTF and PEG, the concentration of rh-CNTF and the pH value were the main influential factors. The optimum pH was 6.5 and the optimal concentration of rh-CNTF was 1 ~ 2 mg / ml. Good quality ratio of 1: 1, time and temperature have little effect. In vitro neurotrophic activity of rh-CNTF was about twice that of PEG-rhCNTF. Conclusion This method optimizes the conditions for the modification of recombinant human ciliary neurons.