Comparison of the RNA Quality of Native Tissue Samples with Tissue Samples after UV Laser Microdissection

To compare untreated tissue samples with fixed and stained tissue samples, three cryo-asserved murine brains of known RNA integrity (RIN) were used. After cryo-asservation at –80 °C, these were transferred to a cryomicrotome and incubated at –35 °C for 45 min. Then the tissue was equilibrated at –18 °C for 30 min and cut into 12 µm sections at –18 °C by cryosectioning. The murine brain sections were divided into two groups which were treated alternately. One (the control group) was directly frozen at –80 °C. The other group was applied to an RNase-free, UV-treated PEN slide. This was done by briefly thawing the sections at room temperature and fixing them for 2 min in 75 % ethanol cooled to –20 °C. After fixation, they were immediately stained with drops of sterile-filtered cresyl violet (CV) dye solution (1 % cresyl violet in 100 % ethanol) and incubated for 45 sec with slight to and fro movement. After this staining procedure, the dye solution was briefly drained off and the slide was immersed in an ascending sequence of ethanol concentrations (75 %, 95 % and 100 %) for 4 sec each and then finally fixed in 100 % ethanol (test group). Following this fixing and staining procedure, the slide was dried in a drying chamber with silica gel for 45 min at room temperature and then stored at –80 °C in the dry box pending laser microdissection.

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