Increased Sensitivity for Determination of Polycyclic Aromatic Hydrocarbon-DNA Adducts in Human DNA Samples by Dissociation-enhanced Lanthanide Fluoroimmunoassay (DELFIA)

Bernadette Schoket, W. Alan Doty, István Vincze, Paul T. Strickland, Giovanni M. Ferri, Giorgio Assennato, Miriam C. Poirier

Research output: Contribution to journalArticlepeer-review

Abstract

A competitive enzyme-linked immunosorbent assay (ELISA), the most frequently used immunoassay for the determination of polycyclic aromatic hydrocarbon- DNA adducts in human tissues, has been modified to achieve approximately a 6-fold increase in sensitivity. The new assay, a competitive dissociation-enhanced lanthanide fluoroimmunoassay (DELFIA) has utilized the same rabbit antiserum as the ELISA, antiserum elicited against DNA modified with benzo[a]pyrene. However, the alkaline phosphatase conjugate has been replaced with a biotin-europium-labeled streptavidin signal amplification system, and the release of europium into the solution forms a highly fluorescent chelate complex that is measured by time-resolved fluorometry. The DELFIA has achieved a 5- to 6-fold increase in sensitivity for measurement of DNA samples modified in vitro with benzo[a]pyrene, for cultured cells exposed to radiolabeled benzo[a]pyrene, and for human samples from occupationally exposed workers. The assay has been validated by comparison of adduct levels determined by DELFIA, ELISA, and radioactivity in DNA from mouse keratinocytes exposed to radiolabeled benzo[a]pyrene. Human lymphocyte DNA samples from 104 Hungarian aluminum plant workers were assayed by ELISA and compared to blood cell DNA samples from 69 Italian coke oven workers assayed by DELFIA. The standard curves demonstrated that the limit of detection of 4.0 adducts in 108 nucleotides for polycyclic aromatic hydrocarbon-DNA adducts by ELISA, using 35 μg of DNA/microtiter plate well, has been decreased to 1.3 adducts in 108 nucleotides by DELFIA, using 20 μg of.

Original languageEnglish (US)
Pages (from-to)349-353
Number of pages5
JournalCancer Epidemiology Biomarkers and Prevention
Volume2
Issue number4
StatePublished - Jul 1 1993

ASJC Scopus subject areas

  • Epidemiology
  • Oncology

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