Jamur Maria Célia, Oliver Constance
Department of Cell and Molecular Biology, Faculdade de Medicina de Ribeirão Preto, University of São Paulo, Ribeirão Preto, SP, Brazil.
Methods Mol Biol. 2010;588:55-61. doi: 10.1007/978-1-59745-324-0_8.
Fixation is one of the most critical steps in immunostaining. The object of fixation is to achieve good morphological preservation, while at the same time preserving antigenicity. Tissue blocks, sections, cell cultures or smears are usually immersed in a fixative solution, while in other situations, whole body perfusion of experimental animals is preferable. Fixation can be accomplished by either chemical or physical methods. The chemical methods include cross-linking agents such as formaldehyde, glutaraldehyde and succinimide esters as well as solvents such as acetone and methanol, which precipitate proteins. Of the physical methods, freezing tissue and air drying are most widely used. This chapter deals with the chemical fixation methods most commonly used for light microscopy.
固定是免疫染色中最关键的步骤之一。固定的目的是在保持良好形态学保存的同时,保留抗原性。组织块、切片、细胞培养物或涂片通常浸入固定液中,而在其他情况下,对实验动物进行全身灌注则更为可取。固定可以通过化学或物理方法来完成。化学方法包括交联剂,如甲醛、戊二醛和琥珀酰亚胺酯,以及沉淀蛋白质的溶剂,如丙酮和甲醇。在物理方法中,冷冻组织和空气干燥应用最为广泛。本章讨论光学显微镜最常用的化学固定方法。