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Detection of Multiple Proteins in Intracerebral Vessels by Confocal Microscopy

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The Blood-Brain and Other Neural Barriers

Part of the book series: Methods in Molecular Biology ((MIMB,volume 686))

Abstract

Assessment of the blood-brain barrier (BBB) may involve the localization of endothelial proteins within the context of endothelial permeability to plasma proteins. The use of antibodies conjugated to fluorescent dyes, coupled with analysis by confocal microscopy, allows for the detection of multiple proteins in components of the neurovascular unit including endothelium and astrocytes. This chapter provides a detailed protocol for detection of three proteins in fixed or frozen sections of rat brain using three fluorophores with unique excitation/emission spectra. Also included is a protocol for tyramide signal amplification, which is useful for detecting proteins of low abundance, and methods for quantitation of intracerebral vessels expressing a particular protein of interest with and without BBB breakdown to plasma proteins.

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Acknowledgements

This work was supported by Grants from the Heart & Stroke Foundation of Ontario #5347, #6003.

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Authors and Affiliations

  1. Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, Canada

    Janet L. Manias, Anish Kapadia & Sukriti Nag

Authors
  1. Janet L. Manias
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  2. Anish Kapadia
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  3. Sukriti Nag
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Editors and Affiliations

  1. Banting Institute, Dept. Laboratory Med. & Pathobiology, University of Toronto, College St. 100, Toronto, M5G 1L5, Ontario, Canada

    Sukriti Nag

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Manias, J.L., Kapadia, A., Nag, S. (2011). Detection of Multiple Proteins in Intracerebral Vessels by Confocal Microscopy. In: Nag, S. (eds) The Blood-Brain and Other Neural Barriers. Methods in Molecular Biology, vol 686. Humana Press. https://doi.org/10.1007/978-1-60761-938-3_7

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  • DOI: https://doi.org/10.1007/978-1-60761-938-3_7

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  • Publisher Name: Humana Press

  • Print ISBN: 978-1-60761-937-6

  • Online ISBN: 978-1-60761-938-3

  • eBook Packages: Springer Protocols

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