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Journal of Histochemistry and Cytochemistry, Vol. 49, 1509-1518, December 2001, Copyright © 2001, The Histochemical Society, Inc.

ARTICLE

Novel Non-isotopic Method for the Localization of Receptors in Tissue Sections

Luc Desnoyersa, Rebecca A. Simonettea, Richard L. Vandlenb, and Brian M. Fendlyc
a Departments of Cell Biology and Technology, Genentech Inc., 1 DNA Way, South San Francisco, California
b Protein Chemistry, Genentech Inc., 1 DNA Way, South San Francisco, California
c Antibody Technology, Genentech Inc., 1 DNA Way, South San Francisco, California

Correspondence to: Luc Desnoyers, Genentech, Inc., 1 DNA Way (m/s 50), South San Francisco, CA 94080. E-mail:desnoyer@gene.com

We describe a novel fluorescent method for the detection of receptors for chimeric proteins in tissue sections. The technique was developed using a recombinant human insulin-like growth factor (IGF-1) chimera, bearing six additional histidine residues at the carboxy-terminal end (IGF-1-His). We demonstrated that dehydration of the tissue sections was detrimental for binding and that its prevention dramatically increased sensitivity. The specificity of IGF-1-His interaction was shown by gradual abolition of the fluorescent signal in the presence of increasing concentrations of IGF-1. Combining immunofluorescence with in situ ligand binding, we showed that IGF-1-His binding corresponded to the IGF-1 receptor (IGFR-1) distribution in human fetal kidney. Moreover, incubation of the tissue sections with an anti-IGFR-1 blocking antibody abolished IGF-1-His binding, demonstrating that the interaction was mediated by the IGFR-1. The method was also used to localize the IGFR-1 in E18 rat embryo sagittal sections. The IGF-1-His binding pattern was observed in brain, cartilage, lung, skin, heart, diaphragm, and tongue, and paralleled the previously reported IGFR-1 distribution. We believe that this new non-isotopic in situ ligand binding method will facilitate rapid and accurate localization of receptors in tissue sections. (J Histochem Cytochem 49:1509–1517, 2001)

Key Words: insulin-like growth factor-1, insulin-like growth factor, receptor-1, ligand binding, receptor, immunofluorescence


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