重组人心肌肌钙蛋白I纯化方法的对比  

Comparison in purification methods of the recombinant human cardiac troponin I

作　　者：郝庆钦[1,2] 周建平[3] 许秀丽[1,2] 刘培[1,2] 温新宇[1] 王玲[1] 田亚平[1] 

机构地区：[1]中国人民解放军总医院生化科,北京100853 [2]南开大学医学院,天津300071 [3]军事医学科学院毒物药物研究所,北京100850

出　　处：《国际检验医学杂志》2014年第14期1817-1818,1820,共3页International Journal of Laboratory Medicine

基　　金：国家高技术研究发展计划(863计划)资助项目(2011AA02A111)

摘　　要：目的对比两种重组人心肌肌钙蛋白I(rhcTnI)纯化方法,获取稳定的rhcTnI,促进心肌肌钙蛋白(cTnI)诊断标准化的研究。方法超声破碎工程菌获取rhcTnI包涵体,经2%Tritonx-100,2mol/L脲洗涤后溶解在8mol/L脲中,分别经CMFF柱上复性和稀释复性纯化rhcTnI,对比两种方法纯化rhcTnI的获得率及其产物在4、-20、-80℃及冻干条件下的稳定性,确立高效获取稳定的cTnI的纯化方法。结果 0.1g湿重包涵体经CM-FF柱上复性和稀释复性的获得率分别为26.8%和18.9%。4、-20、-80℃及冻干条件下,CM-FF下柱上复性获得的rhcTnI稳定,并且柱上复性纯化周期短,效率高。结论 CMFF柱上复性要比稀释复性纯化rhcTnI高效、稳定。Objective To compare the two kinds of purification method for purifying recombinant human cardiac troponin I（cT-nI）to obtain the stable cTnI and promote the study of cTnI diagnosis standardization.Methods The cTnI inclusion body was ob-tained by the ultrasonic broken engineering,after washing by 2% Tritonx-100,2M urea,dissolved in 8M urea,then purified by the column refolding on CM-FF and the dilution refolding respectively.The cTnI yields were compared between the two kinds of meth-od and the stability at 4 ℃,20 ℃,-80 ℃ and on the freeze-dried condition was compared.Then the purification method to effi-ciently obtain the stable cTnI was established.Results The protein about 2 mg and 1.4 mg could be obtained by CM-FF on the col-umn refolding and the dilution refolding from 0.1 g of wet inclusion body,respectively.The former method had the short cycle and high efficiency.The cTnI purified by the column refolding on CM-FF was more stable at 4 ℃,20 ℃,-80 ℃ and on the freeze-dried condition.Conclusion The column refolding on CM-FF is more stable and highly efficient in purification of cTnI than the dilution refolding.

关 键 词：心肌肌钙蛋白I 稀释复性 柱上复性 纯化 

分 类 号：R3411[医药卫生—基础医学]