Journal article

A protocol for cryoembedding the adult guinea pig cochlea for fluorescence immunohistology

Bryony Coleman, Natalie A Rickard, Michelle G de Silva, Robert K Shepherd

JOURNAL OF NEUROSCIENCE METHODS | ELSEVIER SCIENCE BV | Published : 2009

Abstract

Green fluorescent protein (GFP) has been used extensively to label cells in vitro and to track them following their transplantation in vivo. During our studies using the mouse embryonic stem cell line R1 B5-EGFP, we observed variable levels of fluorescence intensity of the GFP within these transfected cells. The variable fluorescence of this protein coupled with the innately autofluorescent nature of several structures within the cochlea collectively made the in vivo identification of these transplanted stem cells difficult. We have modified previously published protocols to enable the discrimination of an authentic GFP signal from autofluorescence in the adult guinea pig cochlea using fluor..

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Grants

Awarded by National Institutes of Health


Funding Acknowledgements

The authors thank the Department of Otolaryngology, University of Melbourne, the National Institutes of Health (Contract NIH-N01-DC-3-1005) and the Bionic Ear Institute for their generous financial support of this project. We thank Dr Steven Backhouse for his assistance with the surgeries and Mrs Maria Clarke for histological advice. BC is supported by a Wagstaff Fellowship in Otolaryngology from the Royal Victorian Eye and Ear Hospital.