不同方法提取肝细胞质膜蛋白效率的比较

Comparison of different methods for extracting plasma membrane protein of liver cell

目的 比较两种提取方法对肝细胞质膜蛋白提取效率的差异,以提高肝细胞质膜蛋白的提取效率.方法 取用手术切取的47例肝细胞癌和43例正常肝组织,分别利用试剂盒和改良法提取肝细胞质蛋白和膜蛋白;BCA法进行蛋白定量后,通过Western blot检测提取出的质蛋白和膜蛋白样品中Na＋/K＋-ATPase和GAPDH相对含量,比较膜蛋白提取特异性.结果 利用改良提取法（B组）对正常肝细胞和肝癌细胞质膜蛋白的提取量分别为（1367.30±486.37）mg/L和（1059.80±226.29）mg/L,均明显高于试剂盒提取量（345.58±117.89）mg/L和（486.24±154.96）mg/L,P＜0.05）;改良提取法提取蛋白中反映质蛋白的指标GAPDH含量显著低于试剂盒提取法（P＜0.05）.结论 通过改良提取方法可有效提高肝细胞膜蛋白提取的丰度和特异性.

Objective To compare two extraction methods in extracting membrane protein of liver cell for improving membrane protein extraction efficiency.Methods Resected tissue from 47 cases of hepatocellular carcinoma （HCC） and 43 cases of normal liver were used to extract protein with extraction kit（A） and improved extraction method（B） ,protein quantitation was evaluated with BCA method,Na＋/K＋ -ATPase and GAPDH were detected between cytoplasmic protein sample and membrane protein sample to display the mixing level between them which was also the extraction specific of membrane protein.Results Liver cell plasma membrane protein extracted in group B was significantly higher than in group A [normal liver：（ 1367.30 ±486.37） mg/L vs （345.58 ± 117.89） mg/L,HCC：（ 1059.80 ± 226.29 ） mg/L vs （486.24 ± 154.96） mg/L,P〈0.05 respectively）.Content of GAPDH in membrane protein extracted by improved extraction method was significantly less than that extracted by extraction kit （ P〈0.05 ）.Conclusion The abundance and specificity of hepatocellular membrane protein extracted with improved extraction method was higher than with extraction kit respectively,which may improve the efficiency of membrane protein research.