Development of monoclonal antibodies specific for 1,N2-ethenodeoxyguanosine and N2,3-ethenodeoxyguanosine and their use for quantitation of adducts in G12 cells exposed to chloroacetaldehyde.
Monoclonal antibodies specific for N2,3-ethenodeoxyguanosine (N2,3-epsilon dGuo) and 1,N2-ethenodeoxyguanosine (1,N2-epsilon dGuo) were developed. In a competitive ELISA, 50% inhibition of binding of the N2,3-epsilon dGuo specific antibody (ETH1) was achieved with 18 fmol of N2,3-epsilon dGuo. Fifty per cent inhibition of the 1,N2-epsilon dGuo-specific antibody (ETH2) required 11 pmol 1,N2-epsilon dGuo. Immunoassays for N2,3-epsilon dGuo and 1,N2-epsilon dGuo in single-stranded DNA were developed using these antibodies. The immunoassays could detect as little as 48 fmol of N2,3-epsilon dGuo or 340 fmol 1,N2-epsilon dGUO in 25 micrograms of single stranded DNA. These assays and previously developed immunoassays for 1,N6-ethenodeoxy-adenosine (1,N6-epsilon dAdo) and 3,N4-ethenodeoxycytidine (3,N4-epsilon dCyd) were used to measure etheno adduct levels in DNA of cells exposed to chloroacetaldehyde. The cells used were V79 cells with an inactivated hprt gene and a single copy of the bacterial gpt gene (G12 cells). The most abundant etheno adduct was 1,N6-epsilon dAdo, followed by 3,N4-epsilon dCyd and N2,3-epsilon dGuo. 1,N2-epsilon dGuo was not detected in chloro-acetaldehyde-treated G12 cells. Chloroacetaldehyde was also shown to be mutagenic in these same cells.
Foiles
P G
PG
Division of Chemical Carcinogenesis, American Health Foundation, Valhalla, NY 10595.
Miglietta
L M
LM
Nishikawa
A
A
Kuśmierek
J T
JT
Singer
B
B
Chung
F L
FL
D000079:Acetaldehyde / Q000031:analogs & derivatives* / Q000633:toxicity; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D004249:DNA Damage; D004277:DNA, Single-Stranded / Q000737:chemistry; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000032:analysis / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D015227:Lipid Peroxidation; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL
D000911:Antibodies, Monoclonal; D004277:DNA, Single-Stranded; C054579:1,N(2)-ethenodeoxyguanosine; C079486:N(2),3-ethenodeoxyguanosine; C004656:chloroacetaldehyde; D003849:Deoxyguanosine; D000079:Acetaldehyde