Poly(ADP-ribose) monoclonal antibody (10H)

The monoclonal antibody 10H is directed against poly(ADP-ribose) (PAR). PAR is synthesized after activation of the nuclear DNA repair enzyme poly(ADP-ribose)polymerase (PARP). PARP is selectively activated by DNA strand breaks to catalyze the addition of long branched chains of PAR to a variety of nuclear proteins, most notably PARP itself.The amount of PAR formed in living cells with DNA damage is commensurate with the extent of the damage. Under DNA damage conditions, PAR undergoes a rapid turnover, with a half-life in the range of minutes, as PAR is rapidly hydrolyzed and converted to free ADP-ribose by the enzyme poly(ADP-ribose)glycohydrolase (PARG). After massive DNA damage (e.g. γ-irradiation or oxidative stress) PAR is detectable in the first 10 minutes and disappears later on. In keratinocytes MAb 10H has been shown to detect UVB-induced apoptosis as early as 4 hour after irradiation, thus being superior to DNA laddering and the TUNEL assay.Due to the very large number of endonuclease-mediated DNA breaks in apoptosis, PARP becomes strongly activated during the so-called execution phase. In the case of DNA damage-induced apoptosis, this represents a "second round" of PAR synthesis. PAR synthesized during apoptosis appears to be remarkably stable. PAR immunofluorescence appears at least as early during apoptosis as does the specific cleavage of PARP by caspase-3. As shown by several groups, this PAR immunofluorescence correlates well with other markers of apoptosis. MAb to Poly(ADP-ribose) (10H) can be used in flow cytometry.A quantitative non-isotopic immuno-dot-blot method for the assessment of cellular poly(ADP-ribosyl)ation capacity using MAb to Poly(ADP-ribose) (10H) has been described.