Jornal Brasileiro de Patologia e Medicina Laboratorial
Print version ISSN 1676-2444
RIBEIRO-SILVA, Alfredo and GARCIA, Sérgio Britto. Comparative study of three different procedures for RNA extraction from formalin-fixed paraffin-embedded samples. J. Bras. Patol. Med. Lab. [online]. 2008, vol.44, n.2, pp. 123-130. ISSN 1676-2444. http://dx.doi.org/10.1590/S1676-24442008000200009.
BACKGROUND: The development of methods for RNA extraction from formalin-fixed paraffin-embedded (FFPE) samples has allowed retrospective studies of molecular biology. OBJECTIVES: To compare the quantity and quality of RNA extracted from FFPE samples using three commercially available kits. Material and methods: Using three different procedures, the total RNA was extracted from 14 paraffin blocks containing fragments of mammary carcinomas, which had been archived for 10 years. The quantity of RNA was expressed in pg/µl; and the quality in RNA integrity number (RIN), by using the Agilent Bioanalyser with Pico LabChip. The RNA with higher RIN extracted from each of the 14 samples was amplified by reverse transcriptase-polymerase chain reaction (RT-PCR) using the G6PD gene with primers designed to create fragments with 67, 151 and 242 base pairs (bp). RESULTS: The mean and median of RNA quantity extracted for the three procedures were respectively 42.91 and 31.31 pg/µl. The mean and median of RIN were respectively 1.8 and 2. In all the samples, the G6PD gene was amplified for RNA fragments with 67 and 151 bp. DISCUSSION: Due to the significant individual variation in quantity and quality of the extracted RNA from each sample, the data from the present study show that, if it is not possible to extract RNA from a given sample in the first attempt, a second extraction should be performed before excluding this sample. CONCLUSION: It was possible to extract RNA with acceptable quality for amplification by RT-PCR (RIN > 1.4) in the three procedures used.
Keywords : RNA; Formalin; Paraffin; Protocol; Molecular biology.