Dube Sukoluhle, Al-Mannai Sharefa, Liu Li, Tomei Sara, Hubrack Satanay, Sherif Shimaa, Jabeen Ayesha, Ahmed Eiman I, Sanchez Apryl, Mifsud William, Bedognetti Davide, Hendrickx Wouter, Raynaud Christophe M
Anatomical Pathology Lab, Department of Pathology, Sidra Medicine, Doha, Qatar.
Tumour Biology and Immunology Laboratory, Research Branch, Sidra Medicine, Doha, Qatar.
J Transl Med. 2025 Jan 6;23(1):11. doi: 10.1186/s12967-024-05890-5.
FFPE tissue samples are commonly used in biomedical research and are a valuable source for next-generation sequencing in oncology, however, extracting RNA from these samples can be difficult the quantity and quality achieved can impact the downstream analysis. This study compared the effectiveness of seven different commercially available RNA extraction kits specifically designed for use with FFPE samples in terms of the quantity and quality of RNA recovered.
This study used 9 samples of FFPE tissue from three different types of tissue (Tonsil, Appendix and lymph node of B-cell lymphoma) to evaluate RNA extraction methods. Three sections of 20 µm of each sample were combined per sample. The slices were distributed in a systematic manner to prevent any biases. Each of the 7 commercially available RNA extraction kits were used according to manufacturer's instructions, with each sample being tested in triplicate resulting in a total of 189 extractions. The concentration, RNA quality score (RQS) and DV200 of each extraction was analysed using a nucleic acid analyser to determine the quantity and quality of the recovered RNA.
This study found that despite processing the FFPE samples in the same standardized way, there were disparities in the quantity and quality of RNA recovered across the different tissue types. Additionally, the study found notable differences in the quantity of RNA recovered when using different extraction kits. In terms of quality, three of the kits performed better than the other four in terms of RQS and DV200 values.
Though many laboratories have developed their own protocols for specific tissue types, using commercially available kits is still a popular option. Although these kits use similar processes and extraction procedures, the amount and quality of RNA obtained can vary greatly between kits. In this study, among the kits tested, while the Roche kit, provided a nearly systematic better-quality recovery than other kits, the ReliaPrep FFPE Total RNA miniprep from Promega yielded the best ratio of both quantity and quality on the tested tissue samples.
福尔马林固定石蜡包埋（FFPE）组织样本常用于生物医学研究，是肿瘤学下一代测序的宝贵来源，然而，从这些样本中提取RNA可能很困难，所获得的数量和质量会影响下游分析。本研究比较了七种专门为FFPE样本设计的市售RNA提取试剂盒在回收RNA的数量和质量方面的有效性。
本研究使用来自三种不同类型组织（扁桃体、阑尾和B细胞淋巴瘤淋巴结）的9个FFPE组织样本评估RNA提取方法。每个样本将三个20μm的切片组合在一起。切片以系统方式分布以防止任何偏差。按照制造商说明使用7种市售RNA提取试剂盒中的每一种，每个样本进行三次测试，总共进行189次提取。使用核酸分析仪分析每次提取的浓度、RNA质量评分（RQS）和DV200，以确定回收RNA的数量和质量。
本研究发现，尽管以相同的标准化方式处理FFPE样本，但不同组织类型回收的RNA在数量和质量上存在差异。此外，研究发现使用不同提取试剂盒时回收的RNA数量存在显著差异。在质量方面，就RQS和DV200值而言，其中三种试剂盒比其他四种表现更好。
尽管许多实验室针对特定组织类型制定了自己的方案，但使用市售试剂盒仍然是一种流行的选择。尽管这些试剂盒使用相似的流程和提取程序，但不同试剂盒获得的RNA数量和质量可能有很大差异。在本研究中，在所测试的试剂盒中，虽然罗氏试剂盒提供的回收质量几乎比其他试剂盒更系统地好，但Promega的ReliaPrep FFPE总RNA微量制备试剂盒在所测试的组织样本上产生了数量和质量的最佳比例。
