This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Zhang, Y.-J. Articles by Santella, R. M. Search for Related Content PubMed PubMed Citation Articles by Zhang, Y.-J. Articles by Santella, R. M.

Quantitation of Aflatoxin B₁-DNA Adducts in Woodchuck Hepatocytes and Rat Liver Tissue by Indirect Immunofluorescence Analysis¹

Comprehensive Cancer Center and Division of Environmental Sciences, School of Public Health, Columbia University, New York, New York 10032 [Y-J. Z., C-J. C., B. H., G-Y. Y., L-L. H., L-W. W., R. M. S.], and Institute of Public Health, National Taiwan University College of Medicine, Taipei, Taiwan 10018 [C-J. C.]

A quantitative indirect immunofluorescence technique was developed utilizing a monoclonal antibody (6A10) recognizing the imidazole ring-opened form of the major N-7 guanine adduct of aflatoxin B₁ (AFB₁). This method was used to investigate adduct formation in woodchuck hepatocytes treated in culture and in liver tissue of rats treated i.p. with AFB₁. Fluorescein isothiocyanate-labeled secondary antiserum was used for adduct localization in conjunction with 4',6-diamidino-2-phenylindole dihydrochloride staining to localize nuclei. Quantitation of AFB₁-DNA adducts was carried out by densitometric analysis of photographic slides. Specific nuclear staining was observed in both woodchuck hepatocytes and rat liver tissue. There was a dose-response relationship between fluorescence intensity and AFB₁ dose in treated animals. Turnover of adducts could also be followed in animals over 48 h with this method. DNA was isolated from liver tissue of treated animals and adduct levels were quantitated by competitive enzyme-linked immunosorbent assay with antibody 6A10 and by fluorescence spectroscopy. There was a significant correlation of the quantitative immunofluorescence intensity with levels of AFB₁ adducts detected by enzyme-linked immunosorbent assay (r = 0.61, P < 0.05) and spectrofluorescence (r = 0.78, P < 0.01). This immunohistochemical method should be applicable to the detection of adducts in liver tissues of humans exposed to high levels of dietary AFB₁.

¹ This study was supported by NIH Grants ES05116 and CA21111 and by agift from the Lucille P. Markey Charitable Trust. Dr. C-J. Chen is the recipientof an NIH Fogarty International Research Fellowship (1 F05 TW04161-01).

² Present address: Department of Biochemistry, Isfahan University of Medical Sciences, Isfahan, Iran.

³ To whom requests for reprints should be addressed, at 650 West 168th Street, New York, NY 10032.

Received 10/ 1/90. Accepted 1/ 3/91.