[Preparation of heat shock protein 70 (Hsp70) and idiotypic determinant single-chain antibody (Id-ScFv) in a patient with B-cell chronic lymphatic leukemia (B-CLL) and antitumor effect of peptide complex Hsp70-Id].

作者：Qin Ye, Zhihua Wang, Shukui Qin · 发表于：PubMed · 年份：2008 · 被引用次数：2 · 研究领域：Heat shock proteins research、Chronic Lymphocytic Leukemia Research、Monoclonal and Polyclonal Antibodies Research

BACKGROUND & OBJECTIVE: Idiotypic determinant (Id) of malignant B lymphocyte surface membrane immunoglobulin (SmIg) is not only a specific marker for chronic B-cell lymphatic leukemia (B-CLL) but also a specific antigen as an attractive target for active immunotherapy. However, as a small self antigen, it has poor immunogenicity. As an important molecule for antigen presentation, heat shock protein 70 (Hsp70) can efficiently strengthen the immunogenicity of antigen. In this study, we prepared Hsp70 and idiotypic determinant single-chain antibody (Id-ScFv) fragment against SmIg in patient with B-CLL and explored the in vitro antitumor effect of peptide complex Hsp70-Id and the possible immune mechanism. METHODS: Id-ScFv and human Hsp70 proteins were expressed in E.coli and identified by sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE) and enzyme-linked immunosorbent assay (ELISA), and purified by metal chelate chromatography and DEAE ion-exchange chromatography. The purified Hsp70 was combined with the prepared Id-ScFv to produce peptide complex Hsp70-Id in vitro. Peripheral blood mononuclear cells (PBMCs) from healthy human were stimulated with Id-ScFv, HSP70 and HSP70-Id, separately. The proliferation rate of PBMCs was determined by MTT assay; the changes of T lymphocyte subsets were assessed by flow cytometry (FCM)û the levels of interleukin-12 (IL-12) and tumor necrosis factor-alpha (TNF-alpha) in supernatant were detected by ELISA. Killing efficiency o...