Multiplex fluorescent in situ hybridization in zebrafish embryos using tyramide signal amplification.

Abstract:

:One of the strengths of the zebrafish is the ease with which in situ hybridization can be performed to determine spatial and temporal patterns of gene expression in whole embryos. Thus far, colorimetric detection methods are mainly used for these analyses. Here we describe a fluorescent in situ hybridization (FISH) protocol for whole-mount zebrafish embryos using tyramide signal amplification (TSA). An optimal set of reagents was identified that allows for simultaneous localization of gene expression patterns of two genes within the same embryo, permitting identification of colocalized expression within single cells. This protocol can be extended to perform multiplex studies by repetition of the TSA-based detection for each target sequentially with a different fluorescent dye label. To this effect, we demonstrate that this approach can be combined with standard horseradish peroxidase (HRP)-mediated immunocytochemistry procedures in addition to FISH.

journal_name

Zebrafish

journal_title

Zebrafish

authors

Clay H,Ramakrishnan L

doi

10.1089/zeb.2005.2.105

subject

Has Abstract

pub_date

2005-01-01 00:00:00

pages

105-11

issue

2

eissn

1545-8547

issn

1557-8542

journal_volume

2

pub_type

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