聚乙二醇修饰重组人干扰素α1b的制备及其性质 |
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引用本文: | 杨宇峰,路煜,石玮,马相虎,秦洁,朱威,楼觉人.聚乙二醇修饰重组人干扰素α1b的制备及其性质[J].粉末涂料与涂装,2010,23(6). |
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作者姓名: | 杨宇峰 路煜 石玮 马相虎 秦洁 朱威 楼觉人 |
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作者单位: | 上海生物制品研究所,上海,200052 |
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基金项目: | 国家863计划资助项目 |
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摘 要: | 目的制备聚乙二醇(PEG)修饰的重组人干扰素α1b(rhIFNα1b),并分析其部分性质。方法采用PEG修饰rhIFNα1b,通过离子交换层析分离纯化修饰混合物,单点修饰产物经超滤浓缩后,经Lowry法检测蛋白的浓度,SDS-PAGE和RP-HPLC检测蛋白的纯度,等电聚焦电泳检测蛋白的等电点,质谱法检测蛋白的相对分子质量,细胞病变抑制法检测蛋白的体外活性,ELISA法检测蛋白的抗原性。结果单点修饰产物的蛋白得率为33%;SDS-PAGE分析纯度为96.1%,RP-HPLC分析纯度为98.1%;等电点为6.22;相对分子质量为39946;比活为1.28×106IU/mg,活性保留率为14.4%;抗原性至少降至原来的1/625。结论制备的单点修饰rhIFNα1b药学性质获得改善,有望开发为安全、长效的新型干扰素。
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关 键 词: | 聚乙二醇 生物 基因修饰 重组人干扰素α 性质 |
Preparation and Properties of PEGylated Recombinant Human Interferon α1b |
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Abstract: | Objective To prepare PEGylated recombinant human interferon α1b(PEG-rhIFNα1b)and analyze its properties.Methods Recombinant human IFNα1b was modified by PEG,and the modified mixture was separated and purified by ion exchange chromatography,based on which PEG-rhIFNα1b was concentrated by ultrafiltration,and determined for concentration by Lowry method,for purity by SDS-PAGE and RP-HPLC,for isoelectric point by isoelectric focusing electrophoresis,for relative molecular mass by mass spectrography,for in vitro activity by cytopathic inhibition assay,and for antigenicity by ELISA.Results The yield of PEGrhIFNα1b was 33%.The purity of PEG-rhIFNα1b was 96.1% by SDS-PAGE and 98.1% by RP-HPLC.The isoelectric point,relative molecular mass,specific activity and activity reservation rate of PEG-rhIFNα1b were 6.22,39 946,1.28 × 106 IU /mg and 14.4% respectively,while the antigenicity decreased by at least 625 folds.Conclusion The pharmacological properties of prepared PEGrhIFNα1b were improved.PEG-rhIFNα1b might be developed to a novel safe and long-acting interferon. |
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Keywords: | PEG Biology gene modification Recombinant human interferon α Property |
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