Chung Joon-Yong, Cho Hanbyoul, Hewitt Stephen M
Experimental Pathology Laboratory, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD.
Department of Obstetrics and Gynecology, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul, Korea.
Biotechniques. 2016 May 1;60(5):239-44. doi: 10.2144/000114415. eCollection 2016.
RNA isolated from formalin-fixed, paraffin-embedded (FFPE) tissue is commonly evaluated in both investigative and diagnostic pathology. However, the quality of the data is directly impacted by RNA quality. The RNA integrity number (RIN), an algorithm based on a combination of electrophoretic features, is widely applied to RNA isolated from paraffin-embedded tissue, but it is a poor indicator of the quality of that RNA. Here we describe the novel paraffin-embedded RNA metric (PERM) for quantifying the quality of RNA from FFPE tissue. The PERM is based on a formula that approximates a weighted area-under-the-curve analysis of an electropherogram of the extracted RNA. Using biochemically degraded RNAs prepared from experimentally fixed mouse kidney specimens, we demonstrate that PERM values correlate with mRNA transcript measurements determined using the QuantiGene system. Furthermore, PERM values correlate with real-time PCR data. Our results demonstrate that the PERM can be used to qualify RNA for different end-point studies and may be a valuable tool for molecular studies using RNA extracted from FFPE tissue.
从福尔马林固定、石蜡包埋(FFPE)组织中分离出的RNA常用于研究性病理学和诊断病理学。然而,数据质量直接受到RNA质量的影响。RNA完整性数(RIN)是一种基于电泳特征组合的算法,广泛应用于从石蜡包埋组织中分离出的RNA,但它并不能很好地指示该RNA的质量。在此,我们描述了一种用于量化FFPE组织RNA质量的新型石蜡包埋RNA指标(PERM)。PERM基于一个公式,该公式近似于对提取RNA的电泳图进行加权曲线下面积分析。使用从实验固定的小鼠肾脏标本制备的生化降解RNA,我们证明PERM值与使用QuantiGene系统测定的mRNA转录本测量值相关。此外,PERM值与实时PCR数据相关。我们的结果表明,PERM可用于鉴定不同终点研究的RNA,并且可能是使用从FFPE组织中提取的RNA进行分子研究的有价值工具。