Development and Validation of a Direct Sandwich Chemiluminescence Immunoassay for Measuring DNA Adducts of Benzo[a]pyrene and Other Polycyclic Aromatic Hydrocarbons

Overview

Journal Mutagenesis

Publisher Oxford University Press

Specialties Environmental Health
Genetics

Date 2012 May 22

PMID 22610669

Citations 5

Authors

Panagiotis Georgiadis

Katalin Kovacs

Stella Kaila

Paraskevi Makedonopoulou

Livia Anna

Miriam C Poirier

Lisbeth E Knudsen

Bernadette Schoket

Soterios A Kyrtopoulos

Affiliations

Soon will be listed here.

Abstract

We have developed and validated a sandwich chemiluminescence immunoassay (SCIA) which measures polycyclic aromatic hydrocarbon (PAH)-DNA adducts combining high throughput and adequate sensitivity, appropriate for evaluation of adduct levels in human population studies. Fragmented DNA is incubated with rabbit antiserum elicited against DNA modified with r7,t8-dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE) and subsequently trapped by goat anti-rabbit IgG bound to a solid surface. Anti-single-stranded (ss) DNA antibodies binds in a quantity proportional to the adduct levels and is detected by chemiluminescence. The BPDE-DNA SCIA has a limit of detection of 3 adducts per 10(9) nucleotides with 5 μg DNA per well. We have validated the BPDE-DNA SCIA using DNA modified in vitro, DNA from benzo[a]pyrene (BP)-exposed cultured cells and mice. The levels of adduct measured by SCIA were lower (30-60%) than levels of bulky DNA adducts measured in the same samples by (32)P-postlabelling. The BPDE-DNA SCIA also detected adducts produced in vivo by PAHs other than BP. When blood DNA samples from maternal/infant pairs were assayed by BPDE-DNA SCIA, the adduct levels obtained were significantly correlated. However, there was no correlation between (32)P-postlabelling and SCIA values for the same samples. The SCIA can be extended to any DNA adduct and is expected to provide, when fully automated, a valuable high-throughput approach in large-scale population studies.

Citing Articles

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Maternal diet during pregnancy and micronuclei frequency in peripheral blood T lymphocytes in mothers and newborns (Rhea cohort, Crete).

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Characterization of solid tumors induced by polycyclic aromatic hydrocarbons in mice.

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Ultrasensitive isolation, identification and quantification of DNA-protein adducts by ELISA-based RADAR assay.

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Micronuclei in cord blood lymphocytes and associations with biomarkers of exposure to carcinogens and hormonally active factors, gene polymorphisms, and gene expression: the NewGeneris cohort.

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