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Originally published as JHC exPRESS on May 3, 2007.
doi:10.1369/jhc.7A7226.2007
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Journal of Histochemistry and Cytochemistry
Volume 55 (9): 925-930, 2007
Copyright ©The Histochemical Society, Inc.

An Improved Method for Three-dimensional Reconstruction of Protein Expression Patterns in Intact Mouse and Chicken Embryos and Organs

Jonas Ahnfelt-Rønne, Mette C. Jørgensen, Jacob Hald, Ole D. Madsen, Palle Serup and Jacob Hecksher-Sørensen

Department of Developmental Biology, Hagedorn Research Institute, Gentofte, Denmark

Correspondence to: Jacob-Hecksher Sørensen, Department of Developmental Biology, Hagedorn Research Institute, Niels Steensens Vej 6, 2820 Gentofte, Denmark. E-mail: jhes{at}hagedorn.dk

We have developed a wholemount immunofluorescence protocol for the simultaneous detection of up to three proteins in mouse and chicken embryos. Combined with Murray's clearing reagent (BABB) and microscope objectives with long working ranges and high numerical apertures mounted on a confocal microscope, cellular resolution can be obtained in depths offering the possibility of examining expression patterns in entire organs or embryos. Three-dimensional projections of the optical confocal sections can be computed with computer software allowing rotation around any axis. The protocol is robust and we find that most antibodies working on tissue sections also work with this protocol. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials. (J Histochem Cytochem 55:925–930, 2007)

Key Words: wholemount • immunofluorescence • confocal • pancreas • three-dimensional • imaging


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