【摘要】 目的:研究Ag85B蛋白促PPD+人外周血单个核细胞的增殖活性。方法:用PCR技术扩增Ag85B基因,构建重组pcDNA3-Ag85B,并将其转染B16细胞株,用RT-PCR鉴定阳性细胞克隆,用Westernblotting鉴定其在阳性细胞克隆内的表达,用3H-TdR掺入法来检测Ag85B的促增殖活性。结果:获得了阳性B16细胞克隆,并鉴定在阳性细胞克隆内有Ag85B成熟蛋白表达;此阳性细胞克隆对PPD+人的PBMC具有较明显地促PBMC增殖活性。结论:转染了pcDNA3-Ag85B的B16阳性细胞克隆对PPD+人的PBMC具有较明显地促PBMC增殖活性,为Ag85B诱发抗肿瘤免疫应答及其机制的研究奠定了基础。更多还原

【Abstract】 Objective: To study the proliferation-activity of Ag85B protein stimulating the PBMCs of PPD⁺ human donors. Methods: The gene encoding Ag85B mature protein was amplified by polymerase chain reaction （PCR）, and recombinant vector （pcDNA₃） was reconstructed and transfected into B16 cell lines. The positive cell clones by RT-PCR and its protein expression was measured by Western blotting. The PBMCs’ proliferation was analyzed by ³[KG*5]H-TdR after the stimulation of positive cell clones in the PBMCs of PPD⁺ and PPD^- human donors. Results: The positive B16 cell clones were obtained and identified by Western blotting the Ag85B mature protein expressed in the clones of positive cells, which could stimulate the PBMCs’ proliferation-activity in PPD⁺ human donors. Conclusion : These modified B16/pcDNA₃-Ag85B cells significantly enhanced the proliferation of PBMCs of PPD⁺ human donors, which provided foundation to the study of Ag85B antitumor immune activity.更多还原