Abstract: ［Abstract］〓Objective〖HTSS〗〓To find a cell lysate suitable for total protein extract from cultured vascular smooth muscle cells for better application in Western blot analysis.
〖HTH〗〖WTHZ〗Methods〖HTSS〗〓Three commonly used cell lysates, solution Ⅰ, solution Ⅱ and solution Ⅲ supplement with protease inhibitors were used to extract total protein from cultured vascular smooth muscle cells. Bradford method was used to determine protein content and SDSpolyacrylamide gel electrophoresis(SDSPAGE) was used to isolate protein. And then, Western blot was used to detect the protein expression of PDGFRβ, Akt, GAPDH and SM22α.
〖HTH〗〖WTHZ〗Results〖HTSS〗〓There was no significant difference in protein concentration among the three cell lysates(P＞005). The optical density values obtaining from cell lysate Ⅱ and Ⅲ were higher than that obtained from cell lysate Ⅰ in small molecular weight proteins(<50 000) detection. The optical density values obtaining from cell lysate Ⅱ and Ⅲ were higher than that obtained from cell lysate Ⅰ, and those values obtaining from cell lysate Ⅲ were higher than that obtained from cell lysate Ⅱ in medium molecular weight proteins(50 000-80 000) detection. The optical density values obtaining from cell lysate Ⅲ were higher than that obtained from cell lysate Ⅰ in high molecular weight protein(>80 000) detection. For Western blot, the band optical density of PDGFRβ detected by cell lysate Ⅱ and Ⅲ was significantly higher than that obtained from cell lysate Ⅰ. The band optical density of Akt detected by cell lysate Ⅱ and Ⅲ was significantly higher than that obtained from cell lysate Ⅰ, and the band optical density of Akt detected from cell lysate Ⅲ was significantly higher than that obtained from cell lysate Ⅱ. The band optical density of GAPDH detected from cell lysate Ⅱ was significantly higher than that obtained from cell lysate Ⅰ and Ⅲ, the band optical density of SM22α detected fromcell lysate Ⅱ was significantly higher than that obtained from cell lysate Ⅰ and Ⅲ(P＜005).
〖HTH〗〖WTHZ〗Conclusion〖HTSS〗〓An optimal cell lysate was selected to detect the different molecular weight protein from cultured vascular smooth muscle cells, which could provide a suitable protocol for the study of protein function based on Western blot assay.

Key words: myocytes, smooth muscle, cell culture techniques, cell lysatessolution