Fixation and staining of LCM slides was conducted under RNAse-free conditions in 50 ml conical polypropylene tubes (Falcon, Franklin Lakes, NJ, USA) containing 45 ml of solution, as previously described [11 (link)]. ProtectRNA™RNAse inhibitor (1:500) (Sigma, Saint Louis, MO, USA) was incorporated in the LCM staining protocol (Table 1 , S3 Fig ) to protect RNA from degradation during water containing steps performed at room temperature (RT) [15 (link)].
LCM slides were moved on dry ice from a slide box into fixative that was prechilled on dry ice for 1 hour (fixative reached a temperature of -20°C). In the OCT removal step (Table 1 ), the solution was applied to sections twice, and the slides were drained on Kimwipes between applications. “One-step Cresyl Violet Acetate / Eosin Y” stain [11 (link)], was modified as follows: 75 μl of cresyl violet stock solution, 25 μl of eosin Y, 250 μl of RNAse-free water and 250 μl of 100% ethanol.
LCM slides were moved on dry ice from a slide box into fixative that was prechilled on dry ice for 1 hour (fixative reached a temperature of -20°C). In the OCT removal step (
Free full text: Click here
