( ly thawed and treated ...)Your Data: RNAlater Stories

Excess RNAlater is removed with a micropipette.

The protocol allows wet sections to be left at room temperature for several days or stored at -20C for longer periods. Treated sections can be shipped at ambient temperature allowing molecular diagnostics to be performed separately from routine diagnostics.

The protocol was tested on numerous breast, prostate, and lung cancer samples. Total RNA (29 g) was regularly recovered depending on the size of the tumor cryosection. RNA isolated by this procedure was consistently of good quality (Figure 1) and of sufficient yield for subsequent qRT-PCR analysis. qRT-PCR was performed on nanogram or smaller quantities of RNA and the recovered RNA was sufficient for measuring the expression levels of hundreds of genes. qRT-PCR results for several genes are shown in Figure 2.

Figure 1. RNA Quality After Isolation from Treated Cryosections. Four 20 m thick sections of prostate, breast and lung tumor material were either snap frozen (lanes labeled snap frozen) or treated with RNAlater as described in the protocol (lanes labeled RNAlater). Alternatively, sections were homogenized in lysis buffer (lanes labeled LB) or transferred to an RNase free microscopic slide, fixed with cold 70% acetone for 2 x 5 min and air dried (lanes labeled acetone). After three days at room temperature, sections were homogenized and RNA was isolated (RNeasy; Qiagen). RNA was separated on an Agilent 2100 bioanalyzer. Shown are gel-like pictures of
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