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Titolo:
ANTIBODIES TO OXIDATIVE DNA-DAMAGE - CHARACTERIZATION OF ANTIBODIES TO 8-OXOPURINES
Autore:
IDE H; KOW YW; CHEN BX; ERLANGER BF; WALLACE SS;
Indirizzi:
UNIV VERMONT,DEPT MICROBIOL & MOL GENET,MARKEY CTR MOL GEN,STAFFORD HALL BURLINGTON VT 05405 UNIV VERMONT,DEPT MICROBIOL & MOL GENET,MARKEY CTR MOL GEN BURLINGTONVT 05405 COLUMBIA UNIV,COLL PHYS & SURG,DEPT MICROBIOL NEW YORK NY 00000
Titolo Testata:
Cell biology and toxicology
fascicolo: 6, volume: 13, anno: 1997,
pagine: 405 - 417
SICI:
0742-2091(1997)13:6<405:ATOD-C>2.0.ZU;2-M
Fonte:
ISI
Lingua:
ENG
Soggetto:
COLI MUTY GENE; ESCHERICHIA-COLI; THYMINE GLYCOL; MONOCLONAL-ANTIBODY; 8-HYDROXYGUANINE 7,8-DIHYDRO-8-OXOGUANINE; URINARY 8-HYDROXY-2'-DEOXYGUANOSINE; MUTAGENIC SUBSTRATE; IONIZING-RADIATION; CRYSTAL-STRUCTURE; OXIDIZED DNA;
Keywords:
DNA ANTIBODIES; 8-OXOADO; 8-OXOGUO; OXIDATIVE DNA DAMAGE;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Science Citation Index Expanded
Citazioni:
59
Recensione:
Indirizzi per estratti:
Citazione:
H. Ide et al., "ANTIBODIES TO OXIDATIVE DNA-DAMAGE - CHARACTERIZATION OF ANTIBODIES TO 8-OXOPURINES", Cell biology and toxicology, 13(6), 1997, pp. 405-417

Abstract

The 8-oxo-7,8-dihydropurines (8-oxopurines) are important cellular premutagenic lesions produced in DNA by free radicals. Specific antibodies were prepared to detect these lesions. For antigens, 8-oxo-7,8-dihydroadenosine (8-oxoAdo) and 8-oxo-7,8-dihydroguanosine (8-oxoGuo) weresynthesized from the bromonucleosides, and the immunogens were produced by conjugating these to either bovine serum albumin or rabbit serumalbumin by the periodate method. Polyclonal antibodies specific for the haptens were elicited from rabbits immunized with the BSA conjugates. The antibodies to 8-oxoAdo (anti-8-oxoAdo) and 8-oxoGuo (anti-8-oxoGuo) precipitated the homologous antigens in an Ouchterlony gel diffusion assay and no cross-reactivity was observed toward the normal nucleosides or to the heterologous 8-oxopurine. Specificity was also examined by hapten inhibition of antibody reactivity with the homologous conjugates using ELISA. For anti-8-oxoAdo, the IC50 for 8-oxoAdo was 8 mumol/L and 8-bromoadenosine, guanosine, and inosine did not inhibit, even at concentrations of 1.25 mmol/L. Similarly, the IC50 for anti-8-oxoGuo for 8-oxoGuo was 0.1 mu mol/L. 8-Methoxyguanosine also inhibitedthe reaction but was about 500-fold less effective than the elicitinghapten. Other nucleosides tested did not inhibit at concentrations upto 100 mu mol/L. Both antibodies could easily detect the corresponding damage in x-irradiated f1 DNA at a dose of 7.5 Gy and both antibodies recognized the corresponding lesion in duplex DNA; however, with anti-8-oxoGuo the signal was reduced about 50% compared to single-stranded DNA. In order to determine the exact amount of each lesion produced in irradiated DNA, and to standardize the ELISA signal, both products were measured after alkaline phosphatase digestion of x-irradiated calf thymus DNA using high-pressure liquid chromatography (HPLC) coupled to an electrochemical detector. Anti-8-oxoGuo could detect ten 8-oxoG residues and anti-8-oxoAdo could detect two 8-oxoA residues per 10 000nucleotides. Thus, these antibodies should be useful for the detection and measurement of 8-oxopurines in cellular DNA.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 03/08/20 alle ore 12:41:00