Alkaline phosphatase fusions of ligands or receptors as in situ probes for staining of cells, tissues, and embryos

Abstract

Publisher Summary Polypeptide ligands and their cell surface receptors bind to one another with high affinity and specificity. These biological properties can be exploited to make affinity probes to detect their cognate ligands or receptors. This approach has been applied for decades, using radiolabeled ligands as probes to detect their receptors. More recently, it has also been found that receptor ectodomains can be used as soluble probes to detect their ligands. When producing soluble receptor or ligand affinity probes, it has been common to produce the probe as a fusion protein with a tag. This can make detection and purification procedures much easier. Two tags that are widely used for this purpose are alkaline phosphatase (AP) or the immunoglobulin Fc region. Both of these tags are dimeric, and are expected to produce a fusion protein with a pair of ligand or receptor moieties facing away from the tag in the same direction. This chapter describes the production of alkaline phosphatase (AP) fusion proteins, and also describes in situ procedures in which these affinity probes are used to detect the distribution of cognate ligands or receptors in tissues or cells.