Comparison of Vibratome and Compresstome sectioning of fresh primate lymphoid and genital tissues for in situ MHC-tetramer and immunofluorescence staining.

Hadia M Abdelaal,Peter J Southern,Ryoko Sawahata,Pamela J Skinner,Hyeon O Kim,Reece Wagstaff

doi:10.1186/s12575-014-0012-4

Hadia M Abdelaal, Peter J Southern + Show 4 more

Open Access

https://doi.org/10.1186/s12575-014-0012-4

Copy DOI

Abstract

BackgroundFor decades, the Vibratome served as a standard laboratory resource for sectioning fresh and fixed tissues. In skilled hands, high quality and consistent fresh unfixed tissue sections can be produced using a Vibratome but the sectioning procedure is extremely time consuming. In this study, we conducted a systematic comparison between the Vibratome and a new approach to section fresh unfixed tissues using a Compresstome. We used a Vibratome and a Compresstome to cut fresh unfixed lymphoid and genital non-human primate tissues then used in situ tetramer staining to label virus-specific CD8 T cells and immunofluorescent counter-staining to label B and T cells. We compared the Vibratome and Compresstome in five different sectioning parameters: speed of cutting, chilling capability, specimen stabilization, size of section, and section/staining quality.ResultsOverall, the Compresstome and Vibratome both produced high quality sections from unfixed spleen, lymph node, vagina, cervix, and uterus, and subsequent immunofluorescent staining was equivalent. The Compresstome however, offered distinct advantages; producing sections approximately 5 times faster than the Vibratome, cutting tissue sections more easily, and allowing production of larger sections.ConclusionsA Compresstome can be used to generate fresh unfixed primate lymph node, spleen, vagina, cervix and uterus sections, and is superior to a Vibratome in cutting these fresh tissues.

Highlights

The Vibratome served as a standard laboratory resource for sectioning fresh and fixed tissues
Over the past decade, we have used Vibratomes extensively to section fresh unfixed lymphoid and genital tissues (Figure 1) from over 200 rhesus macaques and performed in situ tetramer staining combined with immunohistochemistry to develop a temporal and spatial understanding of the progression of SIV infection and the distribution of induced SIV-specific CD8 T cells [1,3,4,5,6,7]
We have used the Compresstome to section fresh unfixed tissues (Figure 1) from nearly 50 rhesus macaques for the combined in situ tetramer and immunohistochemistry staining

Summary

Introduction

The Vibratome served as a standard laboratory resource for sectioning fresh and fixed tissues. In addition to genital [8] and lymphoid tissue [9], the Vibratome has been used to cut fresh tissue sections from brain [10,11], liver [12], kidney [13,14] larynx, thyroid and skeletal muscle tissues [15]. Despite these technical achievements, the standard Vibratome sectioning protocol retains two important disadvantages: the actual sectioning procedure is very time consuming and some tissues are difficult to cut. Vagina and cervix tissues are difficult to cut because sometimes the blade catches on the tissue and pulls it out of the mount

Methods

Results

Conclusion