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To prepare and identify a polyclonal antibody against rat myostatin and investigate myostatin expression in the rat atrophic gastrocnemius muscle after tibial nerve crush.
The purified fusion protein was used as antigen to immunize rabbits for the preparation of polyclonal antibody. The polyclonal antibody of the protein was measured by enzyme linked immunosorbent assay (ELISA), western-blot and immunochemistry. Myostatin protein expression levels in normal and atrophic gastrocnemius muscle were detected by western-blot and immunochemistry assays.
The GST-myostatin had a purity of 96% and possessed high titer and specificity. The level of myostatin in gastrocnemius muscle significantly increased one week after tibial nerve crush, reached the peak on day 14, and then returned to normal level on day 28.
We have successfully made antiserum of rat myostatin and found that the expression level of myostatin protein in the gastrocnemius after tibial nerve crush-induced atrophy was time-dependent. This study provides an experimental basis to clarify the possible role of myostatin during skeletal muscle atrophy.
将纯化好的GST-myostatin融合蛋白免疫家췃, 获得抗血清, 经Western blot, 免疫荧光组织化学及ELISA检测抗体特异性及效价; 建立大鼠胫神经볐伤模型, 采用western blot及免疫组织化学法检测胫神经볐伤后不同时间段腓肠肌中myostatin蛋白表达变化。
得到较高纯度的GST-myostatin融合蛋白, 获得췃源的抗myostatin血清, 经western blot、 免疫组织化学及ELISA实验证实所得抗体具有较高的特异性及效价; 证实在胫神经볐伤后, 腓肠肌myostatin蛋白水平迅速上升, 在第14天达到高峰, 随后逐渐下降, 至第28天降至接近正常水平。
成功制备了췃抗大鼠myostatin抗血清, 进一步检测表明了胫神经损伤后腓肠肌中myostatin蛋白的表达具有时间依赖性。 本研究为阐明myostatin在骨骼肌캮缩过程中的作用提供了实验依据。