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Titolo:
Chemiluminescence enzyme immunoassay of 8-oxoguanine in DNA
Autore:
Bruskov, VI; Masalimov, ZK; Usacheva, AM;
Indirizzi:
Russian Acad Sci, Inst Theoret & Expt Biophys, Pushchino 142292, Moscow Region, Russia Russian Acad Sci Pushchino Moscow Region Russia 142292 cow Region, Russia Pushchino State Univ, Pushchino 142492, Moscow Region, Russia Pushchino State Univ Pushchino Moscow Region Russia 142492 Region, Russia
Titolo Testata:
BIOCHEMISTRY-MOSCOW
fascicolo: 7, volume: 64, anno: 1999,
pagine: 803 - 808
SICI:
0006-2979(199907)64:7<803:CEIO8I>2.0.ZU;2-4
Fonte:
ISI
Lingua:
ENG
Soggetto:
PERFORMANCE LIQUID-CHROMATOGRAPHY; ESCHERICHIA-COLI; REACTIVE OXYGEN; ELECTROCHEMICAL DETECTION; DEOXYRIBONUCLEIC-ACID; MASS-SPECTROMETRY; OXIDATIVE DAMAGE; BASE DAMAGE; 8-HYDROXYGUANINE; 8-HYDROXY-2'-DEOXYGUANOSINE;
Keywords:
7,8-dihydro-8-oxoguanine (8-hydroxyguanine); DNA damage; reactive oxygen species; enzyme immunoassay; enhanced chemiluminescence; monoclonal antibodies;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
53
Recensione:
Indirizzi per estratti:
Indirizzo: Bruskov, VI Russian Acad Sci, Inst Theoret & Expt Biophys, Pushchino 142292, Moscow Region, Russia Russian Acad Sci Pushchino Moscow Region Russia 142292 Russia
Citazione:
V.I. Bruskov et al., "Chemiluminescence enzyme immunoassay of 8-oxoguanine in DNA", BIOCHEM-MOS, 64(7), 1999, pp. 803-808

Abstract

A test system has been developed to determine 8-oxoguanine in DNA, the most important biomarker of damage to DNA bases by reactive oxygen species. The system is based on a chemiluminescence enzyme immunoassay with the use ofmonoclonal antibodies (mcAB) against 8-oxoguanine. The test involves several stages: 1) immobilization of DNA on nitrocellulose membrane filters using an efficient technique with preliminary formation of a complex with protamine sulfate; 2) formation of antigen-antibody complexes (mcAB with 8-oxoguanine in DNA) with secondary antibodies and with a peroxidase-antiperoxidase complex (PAP method); 3) detection of increased chemiluminescence in a solution of hydrogen peroxide, luminol, and p-iodophenol. The increased chemiluminescence is determined with a conventional liquid scintillation counterfor measuring beta-radioactivity. The system was tested by determining 8-oxoguanine formation in DNA upon gamma-irradiation and upon photosensitized oxidation of guanine under visible light in the presence of methylene blue. A linear dose dependence of 8-oxoguanine formation in DNA was shown for gamma-irradiation. The radiation-chemical yield of 8-oxoguanine (G = 0.57 molecule per 100 eV) is convenient to use for calibration of the amount of 8-oxoguanine formed under other conditions. The sensitivity of the method permits the detection of several femtomoles of 8-oxoguanine in a 40 mu g sampleof DNA.

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Documento generato il 13/07/20 alle ore 03:55:08