Purification Of HSP70-polypeptide Complexes And Treatment Of Established Colon Carcinoma-bearing Mice By Dendritic Cells Plused With HSP70-PCs

Objective To establish a new method of purifying heat shock protein 70-polypeptide complexes (HSP70-PCs) by fast protein liquid chromatography (FPLC) system and observe the therapeutic effect of dendritic cells plused with HSP70-PCs.Methods Cytosolic HSP70-PCs of heat-induced colon carcinoma cells were purified successively by chromatographic procedures , using both DEAE-Sepharose chromatographic column and ADP-Agarose chromatographic column. The purified HSP70-PCs were quantified with Bradford assay. The molecular weight and the identity of HSP70-PCs was confirmed by SDS-PAGE and Western-blotting. Bone marrow-derived DCs plused with HSP70-PCs were used to immunize BALB/c mice. The response of cytotoxic T lymphocyte (CTL) was detected. The therapeutic effect induced by DCs was observed on tumor weight and survival time.Results A stained protein with a molecular weight of about 70KD was confirmed by Western-blotting to be HSP70-PCs. The final protein recovery was 63.63μg per 1g tumor cell (wet weight). DCs plused with HSP70-PCs could induce the specific cytotoxic activity of CTL prominently. After the DCs were injected, the tumor growth was significantly inhibited and the survival time of tumor-bearing mice were prolonged conspicuously.Conclusions The high-purity HSP70-PCs of mice colon carcinoma cells can be separated and purified by FPLC rapidly and conveniently. DCs plused with HSP70-PCs have an obvious therapeutic effect on the established colon carcinoma-bearing mice.