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

被引：50
作者：
Affar, EB
Duriez, PJ
Shah, RG
Winstall, E
Germain, M
Boucher, C
Bourassa, S
Kirkland, JB
Poirier, GG
机构：
[1] Univ Laval, Poly ADP Ribose Metab Grp, Hlth & Environm Unit, Med Res Ctr,CHUQ, Ste Foy, PQ G1V 4G2, Canada
[2] Univ Laval, Fac Med, Quebec City, PQ G1V 4G2, Canada
[3] Univ Guelph, Dept Human Biol & Nutr Sci, Guelph, ON N1G 2W1, Canada
来源：
BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS | 1999年 / 1428卷 / 2-3期
基金：
英国医学研究理事会;
关键词：
DNA damage; poly(ADP-ribose) polymerase; anti-poly(ADP-ribose) antibody; poly(ADP-ribose) size; N-methyl-N '-nitro-N-nitrosoguanidine;
D O I：
10.1016/S0304-4165(99)00054-9
中图分类号：
Q5 [生物化学]; Q7 [分子生物学];
学科分类号：
071010 ; 081704 ;
摘要：
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 P-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. (C) 1999 Elsevier Science B.V. All rights reserved.
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收藏
页码：137 / 146
页数：10
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