Cellular Localization of Receptors Using Antibodies Visualized by Light and Dual Labeling Confocal Microscopy

Abstract

Immunocytochemistry can be used to visualize the binding of specific site-directed antisera to receptors in tissue sections and permits the precise identification of cell types expressing a particular receptor when viewed using a conventional light microscope or by confocal microscopy. Protocols are also described for the dual labeling of cells in the same section using primary antisera raised in two different species (one to the receptor of interest, the second to an immunogen such as a cell-specific marker or the endogenous ligand) with the corresponding secondary antisera conjugated to different fluorescent dyes.

The technique has a range of applications. Subtypes of receptors can be identified and distinguished prior to the development of selective agonists or antagonists, which is particularly important for mapping orphan receptors, where the identity of the endogenous ligand in not yet known. The deletion of genes encoding receptors, particularly in mice, has emerged as a powerful tool in understanding the role of a specific receptor in physiological processes. Receptor immunocytochemistry can be used to analyze the resulting phenotype in whole body sections of mice without preselection of the tissue to be studied.