Extraction of RNA from Frozen Sections

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Tissue Handling: Note that all unfixed human tissue should be handled as BioSafety Level 2 materials (wear gloves, lab coat, etc). All plasticware (tubes, tips, etc.) should be RNase free and always handled with gloves to avoid contamination of the sample with RNases. In general, it is not necessary to treat plastic ware to eliminate RNase contamination- just keep separate, unopened stocks of tubes and tips for autoclaving that are always handled while wearing gloves. Similarly, all chemicals used for RNA should be dedicated for that purpose and handled with gloves to avoid contamination.


1. Prepare a scintillation vial (Fisher #3-341-13) for each sample containing 1.5 ml of Trizol (Invitrogen#15596-026). Exercise proper laboratory practice when handling Trizol, as it is toxic and corrosive (i.e., wear gloves, lab coat, eye protection). Also label a 14 ml round bottomed polypropylene centrifuge tube (Falcon #2059) for each sample.


2. In a cryostat at -20°C, cut the desired number of sections and place in the scintillation vial containing Trizol. The scintillation vial is used because the large opening allows the sections to be placed directly in Trizol to prevent degradation of the RNA and sticking of the sections to the sides of the tube. The number of sections to be cut can be estimated as follows:


Pre-cut a thin section (5 mm) and H&E stain it to determine if the sample contains sufficient tumor cells (we prefer at least 70% tumor cells). Measure the length and width of the section. The number of sections needed is then estimated as follows:


Length (mm)