Protocol

Perfusion of Brain Tissues with Fixative

This protocol was adapted from "Techniques," Section IV, in Discovering Neurons: The Experimental Basis of Neuroscience (eds. Paul et al.). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 1997.

INTRODUCTION

In many experiments it is necessary to section the brain to determine the location of a treatment (lesion or electrode) or to look at the histology of the brain using various staining techniques. Because the texture of the brain is so soft (often likened to soft cheese), it must be "fixed" before it can be removed from the skull. A fixative is a chemical that cross-links the molecules of the tissue, rendering it hard and preserving the tissue. This protocol describes a method for perfusing the brain with fixative (specifically, it describes how to perfuse a rat brain; slight modifications may be needed for different animals). A relatively simple gravity feed and the pumping mechanism of the heart is used to get fixative into the brain. A cannula is placed in the heart, or directly in the ascending aorta, of a deeply anesthetized animal. Blood is flushed out with saline first, and then with a fixative. The choice of fixative is often important if a specific staining technique is to be used, especially in immunocytochemistry, because the fixative can interfere with the staining sensitivity.