Multispectral imaging and automated laser capture microdissection of human cortical neurons: a quantitative study of CXCR4 expression

Methods Mol Biol. 2013;1013:31-48. doi: 10.1007/978-1-62703-426-5_3.

Abstract

Quantifying protein and RNA expression within specific cell populations in vivo is an essential step in unraveling the complex mechanisms of neurological disease. The challenges associated with studying human brain tissue are commonly compounded by variations in postmortem interval, formalin fixation time, and tissue processing methods among others. The result is a sample population that is inherently heterogeneous, implying the need for reliable protocols that are sensitive to low levels of antigen while minimizing background and nonspecific staining. Here, we describe a single immunohistochemistry protocol on formalin-fixed, paraffin-embedded human cortex which can be adapted to (1) quantify the relative protein expression of the chemokine receptor, CXCR4, using multispectral image or (2) isolate neuronal RNA through automated laser capture microdissection.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Automation, Laboratory
  • Cerebral Cortex / chemistry*
  • Cerebral Cortex / cytology
  • Fixatives
  • Formaldehyde
  • Humans
  • Image Processing, Computer-Assisted
  • Immunohistochemistry*
  • Laser Capture Microdissection*
  • Microscopy*
  • Neurons / chemistry*
  • Paraffin Embedding
  • RNA / isolation & purification
  • Receptors, CXCR4 / analysis*
  • Receptors, CXCR4 / genetics
  • Tissue Fixation

Substances

  • CXCR4 protein, human
  • Fixatives
  • Receptors, CXCR4
  • Formaldehyde
  • RNA