河北医科大学学报
河北醫科大學學報
하북의과대학학보
JOURNAL OF HEBEI MEDICAL UNIVERSITY
2014年
5期
541-545
,共5页
李江超%杨扬%郝卓芳%周晓明%章倩倩%王丽京
李江超%楊颺%郝卓芳%週曉明%章倩倩%王麗京
리강초%양양%학탁방%주효명%장천천%왕려경
石蜡包埋组织%microRNAs%microRNA 提取%乳腺癌
石蠟包埋組織%microRNAs%microRNA 提取%乳腺癌
석사포매조직%microRNAs%microRNA 제취%유선암
paraffin-embedded tissue%microRNA%extraction%breast cancer
目的:探索一种从石蜡包埋组织标本中快速、便捷、低成本的提取miRNAs的方法。方法收集2年内石蜡包埋的人乳腺癌组织和保存6个月的MMTV基因工程小鼠乳腺癌组织石蜡样本,同时选用进口A公司生产的提取石蜡组织miRNAs试剂盒作为对照;通过2种方法提取人和鼠的石蜡切片中miRNAs做对比分析,包括采用凝胶电泳验证,检测miRNA纯度、浓度分析。为了进一步验证其提取效果,采用定量PCR对所提取的总miRNAs进行miRNA-375和miR-218检测。结果①采用本实验方法从石蜡包埋乳腺癌组织中提取的 miRNAs其纯度在1.86~2.21OD之间,其浓度可以满足常规实验要求(最低38.50mg/L);②荧光定量PCR能够检测到miRNA-375和miRNA-218,其拷贝数与对照组比较,2种方法差异无统计学意义( P﹥0.05)。③该方法提取时间短、成本低,成本是试剂盒法的1/10,提取时间至少节省1/3。结论从石蜡包埋组织标本中提取miRNAs方法切实可行,为从石蜡组织样本提取miRNA进行分子诊断和肿瘤相关的研究提供了便利,对采用回顾性石蜡标本研究miRNAs提供了有力的帮助。
目的:探索一種從石蠟包埋組織標本中快速、便捷、低成本的提取miRNAs的方法。方法收集2年內石蠟包埋的人乳腺癌組織和保存6箇月的MMTV基因工程小鼠乳腺癌組織石蠟樣本,同時選用進口A公司生產的提取石蠟組織miRNAs試劑盒作為對照;通過2種方法提取人和鼠的石蠟切片中miRNAs做對比分析,包括採用凝膠電泳驗證,檢測miRNA純度、濃度分析。為瞭進一步驗證其提取效果,採用定量PCR對所提取的總miRNAs進行miRNA-375和miR-218檢測。結果①採用本實驗方法從石蠟包埋乳腺癌組織中提取的 miRNAs其純度在1.86~2.21OD之間,其濃度可以滿足常規實驗要求(最低38.50mg/L);②熒光定量PCR能夠檢測到miRNA-375和miRNA-218,其拷貝數與對照組比較,2種方法差異無統計學意義( P﹥0.05)。③該方法提取時間短、成本低,成本是試劑盒法的1/10,提取時間至少節省1/3。結論從石蠟包埋組織標本中提取miRNAs方法切實可行,為從石蠟組織樣本提取miRNA進行分子診斷和腫瘤相關的研究提供瞭便利,對採用迴顧性石蠟標本研究miRNAs提供瞭有力的幫助。
목적:탐색일충종석사포매조직표본중쾌속、편첩、저성본적제취miRNAs적방법。방법수집2년내석사포매적인유선암조직화보존6개월적MMTV기인공정소서유선암조직석사양본,동시선용진구A공사생산적제취석사조직miRNAs시제합작위대조;통과2충방법제취인화서적석사절편중miRNAs주대비분석,포괄채용응효전영험증,검측miRNA순도、농도분석。위료진일보험증기제취효과,채용정량PCR대소제취적총miRNAs진행miRNA-375화miR-218검측。결과①채용본실험방법종석사포매유선암조직중제취적 miRNAs기순도재1.86~2.21OD지간,기농도가이만족상규실험요구(최저38.50mg/L);②형광정량PCR능구검측도miRNA-375화miRNA-218,기고패수여대조조비교,2충방법차이무통계학의의( P﹥0.05)。③해방법제취시간단、성본저,성본시시제합법적1/10,제취시간지소절성1/3。결론종석사포매조직표본중제취miRNAs방법절실가행,위종석사조직양본제취miRNA진행분자진단화종류상관적연구제공료편리,대채용회고성석사표본연구miRNAs제공료유력적방조。
Objective To look for better and low cost method for extracting microRNAs ( miRNAs)from paraffin-embedded tissue for miRNA diagnosis or other miRNAs assay. Methods The samples were collected from paraffin-embedded human breast cancer tissue stored over two years and MMTV transgenic mouse paraffin-embedded breast tissue over six months. The miRNAs extract paraffin tissue kit served as control. To detect the quality of miRNAs obtained by these two methods,used 2% gel electrophoresis and the OD value to confirm the quality and concentration. Furthermore,verified miRNAs by testing miRNA-375 and miRNA-218 with quantitative PCR. Results ①miRNAs extracted by our method met the experimental requirements( A260/280=1. 86-2. 21OD),and the lowest concentration was 38. 50mg/L.②The quantitative PCR can detect miRNAs and its copies number compared with the control,the difference was no significant(P﹥0. 05).③This method has relatively lower cost,about 1/10 cost of kit method,while the extraction process decreased about 1/3 time. Conclusion The method we optimized for extracted miRNAs from paraffin-embedded tissue is feasible,it provides a convenient experimental method,and a better approach to detect more retrospective specimens of paraffin-embedded tissue samples.