Methods in plant immunolight microscopy

This chapter describes the methods for preparing plant cells and tissues for immunolight microscopy. Immunocytochemistry takes advantage of the immune reaction between antibody and antigen to identify and localize molecules in cells and tissues. Because antibodies can be made to virtually every biological molecule, this method is at the core of modern cell biology and is rapidly advancing with innovations in microscopy and biochemistry. To detect a target molecule with immunolight microscopy, it is necessary to use an antibody conjugated to an easily recognized tag. The most widely used method is to label an antibody either directly or indirectly with a fluorochrome, a chemical that upon excitation at a specific wavelength will fluoresce in another wavelength. The chapter discusses the indirect immunofluorescence localization of cytoskeletal proteins in isolated plant cells and in intact tissues. However, the fundamental technique can be used to localize other target molecules for which antibodies are available—phytochromes, seed storage proteins, extracellular matrix or cell wall constituents, integrin, calmodulin, and centrin. © 1995, Academic Press Inc.