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Probing the CNS Microvascular Endothelium by Immune-Guided Laser-Capture Microdissection Coupled to Quantitative RT-PCR

  • Nivetha Murugesan
  • Jennifer Macdonald
  • Shujun Ge
  • Joel S. PachterEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 755)

Abstract

Laser-capture microdissection (LCM) allows for retrieval of distinct populations of cells from their closely surrounding neighbors in situ. As such, LCM is highly advantageous for investigating gene expression along the central nervous system (CNS) microvascular endothelium, a tissue that shows both ­considerable segmental and regional heterogeneity. Combining immunohistochemical staining of CNS microvascular endothelial cells with immunofluorescent staining of perivascular astrocytes or smooth muscle cells, immune-guided LCM, immuno-LCM, may be coupled to downstream qRT-PCR to probe varied expression of the endothelium along the CNS microvascular tree during health and disease. Immuno-LCM/qRT-PCR has been used to highlight contributions of the respective segments of the CNS microvasculature to the blood–brain barrier (BBB), and can be employed to examine changes in BBB gene expression ­during pathology.

Key words

LCM Endothelial cell BBB Heterogeneity qRT-PCR 

Notes

Acknowledgments

This work was supported by grants RO-1-MH54718 and R21-NS057241 from the National Institutes of Health, and grant PP-1215 from the National Multiple Sclerosis Society to J.S.P.

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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Nivetha Murugesan
    • 1
  • Jennifer Macdonald
    • 1
  • Shujun Ge
    • 1
  • Joel S. Pachter
    • 1
    Email author
  1. 1.Blood-Brain Barrier Laboratory, Department of Cell BiologyUniversity of Connecticut Health CenterFarmingtonUSA

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