Application of TCA/acetone protein extraction in the cataract lens protein extract

Abstract

AIM: To compare the effectiveness of different protein extraction method in the cataract lens protein extract, and look for the best protein extraction method suitable for cataract proteomics research. METHODS: Cataract lens samples were dissolved in lysis buffer and sonication; the samples were extracted by TCA/acetone and protein extraction kit after centrifugation. Lens proteins of the samples were analyzed by two-dimensional electrophoresis(2-DE), gel electrophoresis and staining, image acquisition and image analysis. RESULTS: According to the results obtained from two-dimensional electrophoresis gel image, the molecular weight of protein points were localized at 14-97.4kDa, PI5-9. The molecular weight of high abundance crystallins were localized at 20-31kDa, the molecular weight of lower abundance crystallins were localized at 35-45kDa. The 2-DE image background of cataract lens sample with hyaluronic acid extracted by TCA /acetone was clear, no obvious vertical bands, point of proteins were more clear, and 35 to 40 protein points were detected. CONCLUSION: TCA/acetone extraction method has better purification compared to other methods in human cataract lens proteomics, and provides the reliable two-dimensional polyacrylamide gel Image for human cataract lens proteomics mass spectrometry analysis.

Keywords

• crystallins
• protein extraction
• two-dimensional gel electrophoresis
• TCA/acetone