Obtaining high quality, intact RNA is the first and often the most critical step in performing gene expression analysis.
Typically, in order to isolate high quality RNA, the tissue has to be processed immediately after harvest. Ambion's patented RNAlater™ Tissue Collection: RNA Stabilization Solution makes it possible for researchers to postpone RNAlater® for days, weeks, or even months after tissue collection without sacrificing RNA integrity (Figure 1). Since RNAlater inactivates all cellular enzymes, including RNases, RNA expression profiles can be "frozen" without immediate RNA isolation.
Figure 1. Quality of RNA Isolated From Tissue Stored in RNA later™ Solution. Fresh mouse tissues were dissected and stored in RNA later at 37°C for 1 day, room temperature for 1 week, or 4°C for 1 month. RNA was isolated using TRI Reagent® (MRC) and analyzed using denaturing agarose gel electrophoresis.
How do you use RNAlater?
Figure 2. Northern Blot of RNA Isolated From RNA later™-Preserved Tissue. Mouse tissues were dissected and stored in RNA later as indicated. RNA was purified from equal mass amounts of tissue using TRI Reagent® (MRC). Five µg of each RNA sample was Northern blotted. The blot was hybridized with 10 6 cpm/ml of a high specific activity probe for p53 and 10 6 cpm/ml of a low specific activity probe for GAPDH.
What can RNAlater do for you?
Treating tissue with RNA later can also preserve the tissue architecture, which is essential for histopathological analysis. Ellis et al (2002) found that treating breast tissue cores with RNA later prior to fixation allows optimal pathological interpretation and preservation of important diagnostic information.
- Dunmire V, Wu C, Symmans WF, Zhang W. (2002) Increased yield of total RNA from fine-needle aspirates for use in expression microarray analysis. Biotechniques 33(4): 890-6.
- Ellis M, Davis N, Coop A, Liu M, Schumaker L, Lee RY, Srikanchana R, Russell CG, Singh B, Miller WR, Stearns V, Pennanen M, Tsangaris T, Gallagher A, Liu A, Zwart A, Hayes DF, Lippman ME, Wang Y, Clarke R. (2002) Development and validation of a method for using breast core needle biopsies for gene expression microarray analyses. Clin Cancer Res 8(5): 1155-66.
- Florell SR, Coffin CM, Holden JA, Zimmermann JW, Gerwels JW, Summers BK, Jones DA, Leachman SA. (2001) Preservation of RNA for functional genomic studies: a multidisciplinary tumor bank protocol. Mod Pathol 14(2): 116-28.
- Bachoon DS, Chen F, Hodson RE. (2001) RNA recovery and detection of mRNA by RT-PCR from preserved prokaryotic samples. FEMS Microbiol Lett 201(2): 127-32.
Recently, Dong-Hun Lee and colleagues at the Lindsley F. Kimball Research Institute of the New York Blood Center compared the above two techniques with the use of RNAlater to stabilize plasma samples during transportation (1). Their investigation showed that use of RNA later greatly improved the nucleic acid testing results for RNA viruses such as Hepatitis C Virus (HCV) and Human Immunodeficiency Virus (HIV). These samples were stable for up to 14 days at room temperature or 37ºC. RNA later was not found to be necessary for shipment of samples where only the HBV DNA virus is analyzed.