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JHC exPRESS: First Published November 12, 2007. doi:10.1369/jhc.7A7187.2007
Copyright © Histochemical Society, Inc.

A more recent version of this article appeared on March 1, 2008.
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Articles

Immunohistochemical Markers for Quantitative Studies of Neurons and Glia in Human Neocortex

Lise Lyck 1, Ishar Dalmau 1, John Chemnitz 1, Bente Finsen 1 and Henrik Daa Schrøder 1*

1 Medical Biotechnology Centre (LL,ID,BF), Anatomy and Neurobiology (JC), and Institute of Clinical Research (HDS), University of Southern Denmark, Odense, Denmark

* To whom correspondence should be addressed. E-mail: henrik.daa.schroeder{at}ouh.regionsyddanmark.dk.

Submitted on January 8, 2007
Accepted on 25 October 2007


  Abstract
Reproducible visualisation of neurons and glia in human brain is essential for quantitative studies of the cellular changes in neurological disease. However, immunohistochemistry in human brain specimens is often compromised due to prolonged fixation. To select cell-lineage specific antibodies for quantitative studies of neurons and the major types of glia, we used 29 different antibodies, different epitope retrieval methods and different detection systems to stain tissue arrays of formalin fixed human brain. The screening pointed at CD45/LCA, CD68(KP1), CNPase, GFAP, HLA-DR, Ki67, NeuN, p25{alpha}-antigen, and S100{beta} as candidates for future cell counting purposes, since these markers visualised specific neuronal and glial cell bodies. Yet, significant negative correlation between staining result and formalin fixation was observed by blinded scoring of staining for CD45/LCA, CNPase, GFAP and NeuN in brain specimens fixed by immersion and stored up to 10 years in 4% formalin solution at room temperature, independent of donor sex and post mortem interval. In contrast, improved preservation of NeuN and CNPase staining, and full preservation of GFAP and CD45/LCA staining in tissue fixed by perfusion and stored for up to 3 years in 0.1% paraformaldehyde solution at 4°C, indicated that immunohistochemistry can be performed in well preserved bio bank material.

Key Words: tissue array, heat induced epitope retrieval, stereology, astrocyte, oligodendrocyte, microglia


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