Reliability of miRNA Analysis from Fixed and Paraffin-Embedded Tissues

Abstract

In clinical practice, patients’ tissues are fixed and paraffin-embedded in order to enable histological diagnosis. Nowadays, those tissues are also used for molecular characterization. Formalin is the most used fixative worldwide, and Bouin’s solution in some worldwide institutions. Among molecular targets, micro RNAs (miRNAs), the single-stranded non-coding RNAs comprised of 18 to 24 nucleotides, have been demonstrated to be resistant to fixation and paraffin-embedding processes, with consequent possible application in clinical practice. In the present study, let-7e-5p, miR-423-3p, miR-92a-1-5p, miR-30d-5p, miR-155-5p, miR-200a-3p, and miR-429 were investigated in formalin and matched Bouin’s solution-fixed tissues of high grade serous ovarian cancers by means of real-time and droplet digital PCR (ddPCR). Micro RNAs were detectable and analyzable in both formalin- and Bouin’s-fixed specimens, but on average, higher Ct values and lower copies/µL were found in Bouin’s-fixed samples. Data from formalin-fixed samples correlated significantly for most targets with Bouin’s ones, except for let-7e-5p and miR-155-5p. This study shows that miRNAs are analyzable in both formalin- and Bouin’s-fixed specimens, with the possibility, after proper data normalization, to compare miRNA-based data from formalin-fixed samples to those of Bouin’s-fixed ones.

Reinfection of Transplanted Livers in HCV- and HCV/HIV-Infected Patients Is Characterized by a Different MicroRNA Expression Profile

“…miRNAs are able to regulate many genes simultaneously, making their action on target pathways more incisive [ 18 ]. Additionally, they are well-preserved in FFPE tissues, thus, making miRNA expression analysis feasible and reliable [ 28 , 29 ].…”

Expression of MicroRNAs in Periodontal Disease: A Systematic Review

“…The long RNA chains are more susceptible to generate cross-links with proteins, which makes their isolation and identification complicated. However, the opposite has been shown: the study of small RNAs in paraffin-embedded tissue is viable, maintaining sufficient integrity and quality for their study [ 24 , 25 ].…”

A Novel HPLC-Based Method to Investigate on RNA after Fixation

“…The amount of unmodified NMP was always higher in Bouin’s fixed samples when compared to formalin-fixed ones (96–100% vs. 80–85%) Bouin’s fixative is made up of picric and acetic acids as coagulant components of the fixative and formaldehyde as cross-linking one [ 27 ]. RNA from BS fixed samples resulted more degraded in terms of fragment detection and RT-PCR analyses, indicating a possible ab initio fragmentation due to the low pH of the fixation solution [ 2 , 6 , 27 ]. The modification of nitrogenous bases in BS fixation is likely avoided or reduced by the acidic pH of the solution [ 28 ].…”

“…The 4% aqueous solution of formaldehyde (formalin) is the most widely used fixative in histopathology. However, in the past also Bouin’s solution was also applied in certain hospitals for its ability to preserve some morphological details, such as nuclear conformation [ 2 ]. In recent years, because of the need of higher quality biomolecules, commercial alcohol-based fixatives have been developed, such as RCL2 [ 3 ], FineFix [ 4 ], Paxgene [ 5 ].…”