Paul Carol Ann, Beltz Barbara, Berger-Sweeney Joanne
CSH Protoc. 2008 Apr 1;2008:pdb.prot4802. doi: 10.1101/pdb.prot4802.
INTRODUCTIONIn 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.
引言
在许多实验中，有必要对大脑进行切片，以确定治疗（损伤或电极）的位置，或者使用各种染色技术观察大脑的组织学结构。由于大脑质地非常柔软（常被比作软奶酪），在从颅骨中取出之前必须进行“固定”。固定剂是一种能使组织分子交联的化学物质，使组织变硬并保存组织。本方案描述了一种用固定剂灌注大脑的方法（具体来说，它描述了如何灌注大鼠大脑；不同动物可能需要稍作修改）。利用相对简单的重力灌注和心脏的泵血机制将固定剂输送到大脑中。将插管置于深度麻醉动物的心脏或直接置于升主动脉中。首先用生理盐水冲洗出血液，然后用固定剂冲洗。如果要使用特定的染色技术，尤其是在免疫细胞化学中，固定剂的选择通常很重要，因为固定剂会干扰染色敏感性。
