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Immunological determination and size characterization of poly(ADP-ribose) synthesized in vitro and in vivo

Immunological determination and size characterization of poly(ADP-ribose) synthesized in vitro and in vivo
Immunological determination and size characterization of poly(ADP-ribose) synthesized in vitro and in vivo
Poly(ADP-ribose) polymerase is a DNA break detecting enzyme playing a role in the surveillance of genome integrity. Poly(ADP-ribose) is synthesized rapidly and transiently from beta-NAD in response to DNA damaging agents. In order to study the physiological significance of poly(ADP-ribose) metabolism, we have developed immunological methods which enable us to study endogenous poly(ADP-ribose) without interfering with cell metabolism and integrity. For this purpose, we produced a highly specific polyclonal anti-poly(ADP-ribose) antibody which immunoreacts with polymers and oligomers. In addition to the immunodot blot method recently described by us (Affar et al., Anal. Biochem. 259 (1998) 280-283), other applications were investigated in cells: (i) detection of poly(ADP-ribose) by ELISA; (ii) characterization of poly(ADP-ribose) size using high resolution gel electrophoresis of polymers, followed by its transfer onto a positively charged membrane and detection with anti-poly(ADP-ribose) antibody; (iii) immunocytochemistry and flow cytometry analyses allowing poly(ADP-ribose) study at the level of individual cells.
dna damage, poly(adp-ribose) polymerase, anti-poly(adp-ribose) antibody, poly(adp-ribose) size, n-methyl-n?-nitro-n-nitrosoguanidine
0304-4165
137-146
Affar, El B.
522c2ca6-a213-4386-8d82-d44595e21685
Duriez, Patrick J.
4cf499bc-007a-43b3-b180-d6e5dc3d151b
Shah, Rashmi G.
e3556b7b-351c-4c95-a201-05d6fd4c030b
Winstall, Eric
90903962-2608-41ed-a27f-2fe2b029b8d0
Germain, Marc
cce211bc-4997-4326-ba61-0888f979ae52
Boucher, Claudia
41c5eb41-c111-4423-9355-c5895250a9db
Bourassa, Sylvie
e2b37ae5-a2f9-4d59-9cf1-13234b075f13
Kirkland, James B.
13dbb2a3-b57f-41e0-be21-a4719d57bc72
Poirier, Guy G.
5151b4a5-5657-412d-88c4-e3b9127ac42c
Affar, El B.
522c2ca6-a213-4386-8d82-d44595e21685
Duriez, Patrick J.
4cf499bc-007a-43b3-b180-d6e5dc3d151b
Shah, Rashmi G.
e3556b7b-351c-4c95-a201-05d6fd4c030b
Winstall, Eric
90903962-2608-41ed-a27f-2fe2b029b8d0
Germain, Marc
cce211bc-4997-4326-ba61-0888f979ae52
Boucher, Claudia
41c5eb41-c111-4423-9355-c5895250a9db
Bourassa, Sylvie
e2b37ae5-a2f9-4d59-9cf1-13234b075f13
Kirkland, James B.
13dbb2a3-b57f-41e0-be21-a4719d57bc72
Poirier, Guy G.
5151b4a5-5657-412d-88c4-e3b9127ac42c

Affar, El B., Duriez, Patrick J., Shah, Rashmi G., Winstall, Eric, Germain, Marc, Boucher, Claudia, Bourassa, Sylvie, Kirkland, James B. and Poirier, Guy G. (1999) Immunological determination and size characterization of poly(ADP-ribose) synthesized in vitro and in vivo. Biochimica et Biophysica Acta (BBA) - Biomembranes, 1428 (2-3), 137-146. (doi:10.1016/S0304-4165(99)00054-9). (PMID:10434031)

Record type: Article

Abstract

Poly(ADP-ribose) polymerase is a DNA break detecting enzyme playing a role in the surveillance of genome integrity. Poly(ADP-ribose) is synthesized rapidly and transiently from beta-NAD in response to DNA damaging agents. In order to study the physiological significance of poly(ADP-ribose) metabolism, we have developed immunological methods which enable us to study endogenous poly(ADP-ribose) without interfering with cell metabolism and integrity. For this purpose, we produced a highly specific polyclonal anti-poly(ADP-ribose) antibody which immunoreacts with polymers and oligomers. In addition to the immunodot blot method recently described by us (Affar et al., Anal. Biochem. 259 (1998) 280-283), other applications were investigated in cells: (i) detection of poly(ADP-ribose) by ELISA; (ii) characterization of poly(ADP-ribose) size using high resolution gel electrophoresis of polymers, followed by its transfer onto a positively charged membrane and detection with anti-poly(ADP-ribose) antibody; (iii) immunocytochemistry and flow cytometry analyses allowing poly(ADP-ribose) study at the level of individual cells.

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More information

Published date: 5 August 1999
Keywords: dna damage, poly(adp-ribose) polymerase, anti-poly(adp-ribose) antibody, poly(adp-ribose) size, n-methyl-n?-nitro-n-nitrosoguanidine

Identifiers

Local EPrints ID: 164809
URI: http://eprints.soton.ac.uk/id/eprint/164809
ISSN: 0304-4165
PURE UUID: 9faf53cc-8357-4f78-b95c-08275c4055f0

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Date deposited: 04 Oct 2010 15:20
Last modified: 16 Jul 2019 23:52

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Contributors

Author: El B. Affar
Author: Patrick J. Duriez
Author: Rashmi G. Shah
Author: Eric Winstall
Author: Marc Germain
Author: Claudia Boucher
Author: Sylvie Bourassa
Author: James B. Kirkland
Author: Guy G. Poirier

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