Detecting and quantifying pADPr in vivo

Methods Mol Biol. 2011:780:117-34. doi: 10.1007/978-1-61779-270-0_8.

Abstract

Poly(ADP-ribose) polymerases (PARP) participate in diverse biological processes contributing to cellular homeostasis or exacerbating injury. PARP catalyzes the addition of ADP-ribose molecules (pADPr) to the target proteins, a process termed poly-ADP-ribosylation. Overactivation of PARP, as reflected by increased poly-ADP-ribosylation, accumulation of pADPr-modified proteins or free pADPr, contributes to the depletion of NAD(+) and mitochondrial dysfunction, potentially leading to cell death via apoptosis or necrosis. Since PARP over-activation has been identified as a key contributor to the pathobiology of many diseases, monitoring PARP 1 activation by detecting and quantifying pADPr may provide valuable mechanistic insights as well as facilitating therapeutic drug monitoring for PARP inhibitors.Several non-isotopic immunodetection methods for quantifying pADPr are discussed: western blotting of poly-ADP-ribosylated proteins, cellular localization of pADPr by immunohistochemistry, quantification of pADPr by enzyme-linked immunoassay and small-scale two-dimensional gel electrophoresis.

Publication types

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

MeSH terms

  • Animals
  • Blotting, Western
  • Electrophoresis, Gel, Two-Dimensional
  • Electrophoresis, Polyacrylamide Gel
  • Enzyme-Linked Immunosorbent Assay
  • Humans
  • Immunohistochemistry
  • Mice
  • NAD
  • Poly Adenosine Diphosphate Ribose / analysis
  • Poly Adenosine Diphosphate Ribose / metabolism
  • Poly(ADP-ribose) Polymerases / analysis
  • Poly(ADP-ribose) Polymerases / metabolism*
  • Rats

Substances

  • NAD
  • Poly Adenosine Diphosphate Ribose
  • Poly(ADP-ribose) Polymerases