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ZFIN ID: ZDB-PUB-210305-8
Whole-mount Immunohistochemistry of Adult Zebrafish Retina for Advanced Imaging
Nagashima, M., Hitchcock, P.F.
Date: 2020
Source: Bio-protocol   10: e3848 (Journal)
Registered Authors: Hitchcock, Peter, Nagashima, Mikiko
Keywords: Antibody staining, Microscopy, Müller glia, Photoreceptor, Visual system
MeSH Terms: none
PubMed: 33659497 Full text @ Bio Protoc
ABSTRACT
Immunohistochemistry is a widely used technique to examine the expression and subcellular localization of proteins. This technique relies on the specificity of antibodies and requires adequate penetration of antibodies into tissues. The latter is especially challenging for thick specimens, such as embryos and other whole-mount preparations. Here we describe an improved method of immunohistochemistry for retinal whole-mount preparations. We report that a cocktail of three reagents, Triton X-100, Tween-20, and DMSO, in blocking and antibody dilution buffers strongly enhances immunolabeling in whole-mount retinas from adult zebrafish. In addition, we establish that in whole retinal tissues, a classic epitope retrieval method, based on citrate buffer, is effective for immunolabeling membrane-associated proteins. Overall, this simple modification allows precise and reproducible immunolabeling of proteins in retinal whole-mounts.
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