人舌癌Tca8113细胞热休克蛋白多肽复合物(HSP70-PC)的提取和鉴定

Purification and identification of HSP70 tumor peptide extracted from heated human Tca8113 cells

采用ADP-Agarose亲和层析结合DEAE离子交换对Tca8113细胞的HSP70-PC进行分离纯化。结果显示在NaCl250～350mmol/L浓度处出现一洗脱峰,收集此峰各组分,经10%SDS-PAGE、在相对分子质量70000处可见清晰单一蛋白条带,电泳后于第5峰发现相对分子质量为70000的蛋白,此蛋白被洗脱的盐浓度为NaCl250～350mmol/L,此即为高纯度的HSP70-PC。HSP70获得率为1g湿重的Tca8113细胞能纯化85μg的HSP70-PC。采取低渗匀浆、超速离心、ConA-Sepharose亲和层析、ADP-Agarose亲和层析及DEAE离子交换的纯化方案,可获得高纯度的HSP70-PC。

To study the purification and identification of HSP70 tumor peptide extracted from heated human Tca8113 cells.HSP70 associated Tca8113 cells tumor peptides were purified from tumor cell pellet treated in water bath（43 ℃/30 min）,by ConA-Sepharose column,ADP-Agarose column and DEAE ino-exchange chromatography.The purified fractions were tested by SDS-PAGE,immunoblotted with anti-HSP70 mAb.HSP70 tumor peptide was purified from heated Tca8113 cells,which was a high purity protein with 70 kD molecular weight and integrated with anti-HSP70 mAb.The highly purified HSP70-PC can be obtained successfully by using the affinity chromatography and DEAE ion-exchange chromatography.