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Ice incubation step allows adherence of archival histological specimens on gelatin-coated slides for TUNEL staining
G M Seigel1, J Ambati, W Richard Green
1Department of Neurobiology and Anatomy, University of Rochester, School of Medicine and Dentistry, 601 Elmwood Avenue, Rochester, NY 14642, USA. gail_seigel@urmc.rochester.edu
Abstract:
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.
Insights
A simple ice-incubation step improves tissue adherence on gelatin slides during histological procedures like terminal deoxynucleotidyl transferase nick-end labelling (TUNEL) staining. This method enhances the reliability of archival tissue analysis.
Area of Science:
- Histology and Pathology
- Molecular Biology Techniques
Background:
- Gelatin-coated slides exhibit poor tissue adherence during histological procedures requiring 37°C incubations.
- This issue complicates analyses such as terminal deoxynucleotidyl transferase nick-end labelling (TUNEL).
Purpose of the Study:
- To develop a straightforward method to improve tissue adherence on gelatin-coated slides during 37°C incubations.
- To enable successful analysis of archival ocular tissue sections.
Main Methods:
- Implementing a 30-minute ice-incubation step immediately after the 37°C terminal deoxynucleotide transferase reaction.
- Continuing with the standard TUNEL staining protocol after the ice incubation.
Main Results:
- The ice-incubation step effectively re-solidified melted gelatin, ensuring tissue specimen adherence.
- Successfully analyzed 14 archival specimens using the modified TUNEL staining protocol.
Conclusions:
- A simple post-incubation cooling step significantly enhances tissue adherence on gelatin slides.
- This method has broad applicability for various histological protocols requiring 37°C incubations, including TUNEL, in situ hybridization, and immunohistochemistry.
