Protocol

Cryosectioning Tissues

This protocol was adapted from "Preparation of Cells and Tissues for Fluorescence Microscopy," Chapter 4, in Basic Methods in Microscopy (eds. Spector and Goldman). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 2006.

INTRODUCTION

Cryosections are rapidly and relatively easily prepared prior to fixation, and they provide a good system for visualizing fine details of the cell. Although cryosections are physically less stable than paraffin- or resin-embedded sections, they are generally superior for the preservation of antigenicity and therefore the detection of antigens by microscopy. The preparation of cryosections does not involve the dehydration steps typical of other sectioning methods, and, furthermore, sectioning, labeling, and observation of specimens can usually be carried out in one day. In general, the sample is frozen quickly in either isopentane or liquid nitrogen. (Small samples such as cells and small tissues may be mixed in a slurry of an inert support medium such as optimal cutting temperature [OCT] compound before freezing). Rapid freezing reduces ice crystal formation and minimizes morphological damage. Frozen sections may be used for a variety of procedures, including immunochemistry, enzymatic detection, and in situ hybridization. A protocol for cryosectioning is presented here.