Immunoelectron microscopy of cryofixed and freeze-substituted plant tissues

Abstract

Cryofixation and freeze-substitution techniques preserve plant ultrastructure much better than conventional chemical fixation techniques. The advantage of cryofixation is not only in structural preservation, as seen in the smooth plasma membrane, but also in the speed in arresting cell activity. Immunoelectron microscopy reveals the subcellular localization of molecules within cells. Immunolabeling in combination with cryofixation and freeze-substitution techniques provides more detailed information on the immunoelectron-microscopic localization of molecules in the plant cell than can be obtained from chemically fixed tissues. Here, we introduce methods for immunoelectron microscopy of post-embedded, cryofixed plant tissues by applying an antibody to a thin plastic resin-embedded section prepared by cryofixation followed by freeze-substitution.