What is the most important pretreatment for aging slides for FISH?

The most critical pretreatment for aging slides for Fluorescence In Situ Hybridization (FISH) is the application of 2x SSC (saline sodium citrate) at 37 degrees for 30 minutes. The use of 2x SSC serves to rehydrate and prepare the slides containing fixed cells, which helps to improve the hybridization efficiency of probes to their target sequences on the chromosomes.

Performing this pretreatment at 37 degrees is significant because the moderate heat enhances the penetration of the fixative and helps in the renaturation of the DNA strands, allowing for optimal binding of the FISH probes. The temperature aids in the dissociation and stabilization of nucleic acid interactions, which is crucial for achieving specific and effective hybridization.

Timing is also important; a duration of 30 minutes balances the need for effective treatment while preventing any potential degradation of the DNA due to prolonged exposure to elevated temperature. This preparation step ensures that the chromosomes are in a state conducive to hybridization, which is fundamental for accurate results in FISH analysis.