Ying Hu1, Haibo Han1, Yixue Wang1, Lijie Song1, Xiaojing Cheng2, Xiaofang Xing2, Bin Dong3, Xiaohong Wang1, Meng Chen1, Lianhai Zhang4, Jiafu Ji1,4. 1. 1 Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education), Department of BioBank, Peking University Cancer Hospital and Institute , Beijing, China . 2. 2 Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education), Division of Gastrointestinal Cancer Translational Research Laboratory, Peking University Cancer Hospital and Institute , Beijing, China . 3. 3 Department of Pathology, Peking University Cancer Hospital and Institute , Beijing, China . 4. 4 Department of Gastrointestinal Surgery, Peking University Cancer Hospital and Institute , Beijing, China .
Abstract
BACKGROUND: Comparative analysis of RNA expression profiles between cancer and adjacent noncancerous tissues is an important part of cancer research. High-quality RNA is essential for consistent, reliable results, especially for identification of cancer biomarkers. However, the impact of freeze-thaw cycles on the quality of RNA both in gastrointestinal cancer and paired adjacent tissues is still unclear. AIM: To investigate the influence of freeze-thaw cycles on RNA integrity and overall histomorphology of gastrointestinal cancer and paired adjacent noncancerous tissues. METHODS: Gastrointestinal cancer and matched adjacent noncancerous tissues were frozen and thawed twice before extracting RNA. Total RNA in each sample was extracted with TRIzol reagents and the RNA integrity was assessed by RNA integrity number (RIN) on an Agilent Bioanalyzer. Light microscopy was then used to assess tissue composition and morphology. RESULTS: RIN values for all samples tended to decrease in correlation with the frequency of freeze-thawing. With an RIN cutoff value of 6, RNA extracted from pancreatic cancer tissues was not qualified after the first freeze-thaw cycle. Moreover, all RNA extracted from adjacent noncancerous tissues had nonqualifying RIN scores after the first freeze-thaw cycle, except for liver tissues. Microscopically, all samples displayed qualified tissue morphology regardless of freeze-thaw cycle frequency. CONCLUSION: Freeze-thawing affects the RNA integrity, but not the tissue morphology of gastrointestinal cancer and paired adjacent noncancerous tissues. Furthermore, the RNA extracted from adjacent noncancerous tissues is more easily degraded than that in cancer tissues.
BACKGROUND: Comparative analysis of RNA expression profiles between cancer and adjacent noncancerous tissues is an important part of cancer research. High-quality RNA is essential for consistent, reliable results, especially for identification of cancer biomarkers. However, the impact of freeze-thaw cycles on the quality of RNA both in gastrointestinal cancer and paired adjacent tissues is still unclear. AIM: To investigate the influence of freeze-thaw cycles on RNA integrity and overall histomorphology of gastrointestinal cancer and paired adjacent noncancerous tissues. METHODS:Gastrointestinal cancer and matched adjacent noncancerous tissues were frozen and thawed twice before extracting RNA. Total RNA in each sample was extracted with TRIzol reagents and the RNA integrity was assessed by RNA integrity number (RIN) on an Agilent Bioanalyzer. Light microscopy was then used to assess tissue composition and morphology. RESULTS: RIN values for all samples tended to decrease in correlation with the frequency of freeze-thawing. With an RIN cutoff value of 6, RNA extracted from pancreatic cancer tissues was not qualified after the first freeze-thaw cycle. Moreover, all RNA extracted from adjacent noncancerous tissues had nonqualifying RIN scores after the first freeze-thaw cycle, except for liver tissues. Microscopically, all samples displayed qualified tissue morphology regardless of freeze-thaw cycle frequency. CONCLUSION: Freeze-thawing affects the RNA integrity, but not the tissue morphology of gastrointestinal cancer and paired adjacent noncancerous tissues. Furthermore, the RNA extracted from adjacent noncancerous tissues is more easily degraded than that in cancer tissues.
Entities:
Keywords:
RNA integrity number (RIN); freeze-thaw cycles