Document Detail


Immunostaining of dissected zebrafish embryonic heart.
MedLine Citation:
PMID:  22258109     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Zebrafish embryo becomes a popular in vivo vertebrate model for studying cardiac development and human heart diseases due to its advantageous embryology and genetics. About 100-200 embryos are readily available every week from a single pair of adult fish. The transparent embryos that develop ex utero make them ideal for assessing cardiac defects. The expression of any gene can be manipulated via morpholino technology or RNA injection. Moreover, forward genetic screens have already generated a list of mutants that affect different perspectives of cardiogenesis. Whole mount immunostaining is an important technique in this animal model to reveal the expression pattern of the targeted protein to a particular tissue. However, high resolution images that can reveal cellular or subcellular structures have been difficult, mainly due to the physical location of the heart and the poor penetration of the antibodies. Here, we present a method to address these bottlenecks by dissecting heart first and then conducting the staining process on the surface of a microscope slide. To prevent the loss of small heart samples and to facilitate solution handling, we restricted the heart samples within a circle on the surface of the microscope slides drawn by an immEdge pen. After the staining, the fluorescence signals can be directly observed by a compound microscope. Our new method significantly improves the penetration for antibodies, since a heart from an embryonic fish only consists of few cell layers. High quality images from intact hearts can be obtained within a much reduced procession time for zebrafish embryos aged from day 2 to day 6. Our method can be potentially extended to stain other organs dissected from either zebrafish or other small animals.
Authors:
Jingchun Yang; Xiaolei Xu
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural; Video-Audio Media     Date:  2012-01-10
Journal Detail:
Title:  Journal of visualized experiments : JoVE     Volume:  -     ISSN:  1940-087X     ISO Abbreviation:  J Vis Exp     Publication Date:  2012  
Date Detail:
Created Date:  2012-01-19     Completed Date:  2012-03-21     Revised Date:  2013-12-09    
Medline Journal Info:
Nlm Unique ID:  101313252     Medline TA:  J Vis Exp     Country:  United States    
Other Details:
Languages:  eng     Pagination:  e3510     Citation Subset:  IM    
Copyright Information:
Copyright © 2012 Journal of Visualized Experiments
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MeSH Terms
Descriptor/Qualifier:
Animals
Cardiac Surgical Procedures
Heart / anatomy & histology*,  embryology*
Immunohistochemistry / methods*
Myocardium / cytology,  ultrastructure*
Zebrafish
Grant Support
ID/Acronym/Agency:
HL81753/HL/NHLBI NIH HHS; R01 HL081753/HL/NHLBI NIH HHS; R01 HL107304/HL/NHLBI NIH HHS
Comments/Corrections

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