title: RNA stability in human liver: comparison of different processing times, temperatures and methods.
abstract: The accuracy of information garnered by real-time quantitative polymerase chain reaction (RT-qPCR), an important technology for elucidating molecular mechanisms of disease, is dependent on tissue quality. Thus, this study aimed to determine the effects of intra-operative manipulation, extended processing times, different temperatures or storage in RNAlater on RNA quality in liver samples for tissue banking. Liver samples, flash-frozen or in RNAlater, were collected over a time course (during surgery before blood arrest up to 1 day after surgery) with samples kept either at room temperature (RT) or on ice. This study showed that at the longest time-point at RT, the RNA quality decreased significantly by 20%. However, relative gene expressions of FOS, GUSB, MYC, HIF1α and GFER were in general not significantly different when the time-points were compared. In conclusion, samples should be kept on ice during processing, and either RNAlater or snap-freezing should be utilised for storage. Further, intra-operative manipulation and extended postoperative processing time generally does not change relative gene expression levels for the 5 genes studied, making such sampling suitable for RT-qPCR analysis. Thus, if relative gene expression of a gene of interest is stable, these guidelines will lead to increased accrual of samples to the tissue bank.
authors.affiliation: Grosshadern Hospital Tissue Bank, Department of Surgery, Grosshadern Hospital, Ludwig Maximilians University, Munich, Germany.
authors.forename: Serene M L
authors.initials: SM
authors.lastname: Lee
authors.forename: Celine
authors.initials: C
authors.lastname: Schelcher
authors.forename: Sevdije
authors.initials: S
authors.lastname: Gashi
authors.forename: Stefanie
authors.initials: S
authors.lastname: Schreiber
authors.forename: Reinhard M K
authors.initials: RM
authors.lastname: Thasler
authors.forename: Karl-Walter
authors.initials: KW
authors.lastname: Jauch
authors.forename: Wolfgang E
authors.initials: WE
authors.lastname: Thasler
mesh_terms: D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D005091:Exons; D005615:Freezing; D015870:Gene Expression; D006801:Humans; D008099:Liver / Q000378:metabolism*; D012313:RNA / Q000235:genetics / Q000302:isolation & purification*; D020871:RNA Stability / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D013048:Specimen Handling / Q000379:methods; D013997:Time Factors; D014015:Tissue Banks
chemical_list: D018076:DNA, Complementary; D012313:RNA