Ice incubation step allows adherence of archival histological specimens on gelatin-coated slides for TUNEL staining.

Gelatin-coated slides provide poor tissue adherence during histological procedures which require 37 degreesC incubations, such as terminal deoxynucleotidyl transferase nick-end labelling (TUNEL) analysis. We encountered this difficulty during attempts to analyze archival ocular tissue sections which had been previously sectioned and mounted on gelatin-coated slides. The solution to this problem turned out to be relatively straightforward: Immediately after the 37 degreesC terminal deoxynucleotide transferase step, we incubated the slides on ice for 30 min before continuing with the remainder of the protocol. This ice-incubation step re-solidified the melted gelatin, which allowed for continued adherence of the tissue specimen for further manipulations. This modified TUNEL staining protocol has been used successfully to analyze 14 archival specimens thus far, which would have been nearly impossible to accomplish otherwise. We believe that this ice re-solidification step for gelatin-coated slides has broad applications for procedures which require 37 degreesC incubations, including TUNEL staining, as well as other in situ hybridization and immunohistochemistry protocols.