AIM: We optimized a rapid and efficient tissue lysis method using the MagNA Lyser (Roche, Germany). Using this novel method combined with immunoblot analysis, we investigated the correlation between abnormal Bcl-XL ex...AIM: We optimized a rapid and efficient tissue lysis method using the MagNA Lyser (Roche, Germany). Using this novel method combined with immunoblot analysis, we investigated the correlation between abnormal Bcl-XL expression and clinicopathological characteristics in colorectal cancer. METHODS: Tissue samples from Sprague-Dawley rats were tested to determine optimal lysis conditions for use with MagNA Lyser. We next used the new method to extract tissue proteins from the tumor tissue of a colorectal cancer patient. The availability of extractable tissue proteins for proteomic study was demonstrated by two-dimensional (2D) gel electrophoresis and subsequent matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. In addition, we prepared tissue lysates from paired tumor tissues and adjacent nontumor tissues of 50 colorectal carcinoma patients. Ensuing immunoblot analyses were performed to detect the level of Bcl-XL expression. RESULTS: The optimal sample sizes processed were found to be around 200 mg, with oscillation frequency of 6 500 r/min for 80 s. Test of the first human tissue lysate confirmed that the MagNA Lyser method was adequate for protein extraction and subsequent identification by current proteomic protocols. The method was also applicable to immunoblot analysis. Thirty of 50 (60%) colorectal patients exhibited higher level of Bcl-XL expression in their tumor tissues. Raised level of Bcl-XL expression correlated with patients' gender and tumor cell proliferation index (P= 0.037 and P<0.001, respectively), but was independent of clinicopathological characteristics and overall survival. CONCLUSION: We report a novel tissue lysis method applicable to proteomic and immunoblot analyses, which can facilitate the discovery and detection of cancer protein alterations.展开更多
文摘AIM: We optimized a rapid and efficient tissue lysis method using the MagNA Lyser (Roche, Germany). Using this novel method combined with immunoblot analysis, we investigated the correlation between abnormal Bcl-XL expression and clinicopathological characteristics in colorectal cancer. METHODS: Tissue samples from Sprague-Dawley rats were tested to determine optimal lysis conditions for use with MagNA Lyser. We next used the new method to extract tissue proteins from the tumor tissue of a colorectal cancer patient. The availability of extractable tissue proteins for proteomic study was demonstrated by two-dimensional (2D) gel electrophoresis and subsequent matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. In addition, we prepared tissue lysates from paired tumor tissues and adjacent nontumor tissues of 50 colorectal carcinoma patients. Ensuing immunoblot analyses were performed to detect the level of Bcl-XL expression. RESULTS: The optimal sample sizes processed were found to be around 200 mg, with oscillation frequency of 6 500 r/min for 80 s. Test of the first human tissue lysate confirmed that the MagNA Lyser method was adequate for protein extraction and subsequent identification by current proteomic protocols. The method was also applicable to immunoblot analysis. Thirty of 50 (60%) colorectal patients exhibited higher level of Bcl-XL expression in their tumor tissues. Raised level of Bcl-XL expression correlated with patients' gender and tumor cell proliferation index (P= 0.037 and P<0.001, respectively), but was independent of clinicopathological characteristics and overall survival. CONCLUSION: We report a novel tissue lysis method applicable to proteomic and immunoblot analyses, which can facilitate the discovery and detection of cancer protein alterations.
