人骨肉瘤组织热休克蛋白gp96-肽复合物的提取、纯化与鉴定

Isolation, purification and identification of HSPgp96-peptide complex from human osteosarcom tissues

目的从人骨肉瘤组织中提取、纯化热休克蛋白（HSP）gp96-肽复合物的方法。方法采用三步蛋白纯化法从15例人骨肉瘤患者肿瘤组织中提取、纯化HSPgp96-肽复合物。SDS—PAGE、Western blot鉴定HSPgp96-肽复合物，应用改良的Bradford计算蛋白得率。结果应用该方法提取的蛋白质经SDS—PAGE和Western blot特异性鉴定，证实是HSPgp96-肽复合物；蛋白得率为每g湿重骨肉瘤组织可纯化HSPgp96-肽复合物30～40μg／g。结论三步蛋白纯化法不仅操作简便、重复性高，而且提取的HSPgp96-肽复合物具有高获率、高纯度、高特异性等特点。

Objective To derive and purify HSPgp96-peptide complex from human osteosarcoma tissues. Methods Three-step purification of HSPgp96 and Western-blot were used to derive, purify and certify HSPgp96-peptide complex from 15 osteosarcoma tissues, and advanced Bradford＇ s method was used to calculate the final protein recovery. Results The HSPgp96-peptide complex had a high purity and specificity. The final protein recovery was 30-40 μg/g tumor tissue （wet weight）. Conclusion Three-step purification of HSPgp96 is simple and has a high reproducibility. HSPgp96-peptide complex obtained by this method has the merits of high purity, and high specificity.