Skip to main content
NCBI home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1989 Jan 25;17(2):539–551. doi: 10.1093/nar/17.2.539

Detection of specific DNA sequences using antibodies recognizing UV-labelled DNA.

V Lund 1, B H Lindqvist 1, G Eggset 1
PMCID: PMC331602  PMID: 2464795

Abstract

This non-isotopic method for detection of nucleic acids is based on the in situ labelling of the nucleic acid by exposure to UV-irradiation. The different UV-induced photoproducts, mainly of the thymidine dimer type, are recognized by purified rabbit antibodies specific to the lesions introduced. The UV-labelled nucleic acids can then be visualized by conventional immunostaining procedures. A major advantage of the technique is the low cost and the ease by which the DNA is specifically labelled. The purified rabbit antibodies were shown to be specific for UV-irradiated DNA, and the method was applied for detection of specific DNA sequences hybridized to homologous target DNA on membrane support. We believe that the sensitivity of the method can be improved, and the significance of using different UV-doses, immunostaining methods and membrane types is discussed.

Full text

PDF
539

Images in this article

544Image
on p.544
545Image
on p.545
546Image
on p.546
547Image
on p.547
548Image
on p.548

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Adams J. C. Heavy metal intensification of DAB-based HRP reaction product. J Histochem Cytochem. 1981 Jun;29(6):775–775. doi: 10.1177/29.6.7252134. [DOI] [PubMed] [Google Scholar]
  2. Bittner M., Kupferer P., Morris C. F. Electrophoretic transfer of proteins and nucleic acids from slab gels to diazobenzyloxymethyl cellulose or nitrocellulose sheets. Anal Biochem. 1980 Mar 1;102(2):459–471. doi: 10.1016/0003-2697(80)90182-7. [DOI] [PubMed] [Google Scholar]
  3. Chomczynski P., Qasba P. K. Alkaline transfer of DNA to plastic membrane. Biochem Biophys Res Commun. 1984 Jul 18;122(1):340–344. doi: 10.1016/0006-291x(84)90480-7. [DOI] [PubMed] [Google Scholar]
  4. Eggset G., Volden G., Krokan H. Characterization of antibodies specific for UV-damaged DNA by ELISA. Photochem Photobiol. 1987 Apr;45(4):485–491. doi: 10.1111/j.1751-1097.1987.tb05407.x. [DOI] [PubMed] [Google Scholar]
  5. Eggset G., Volden G., Krokan H. U.v.-induced DNA damage and its repair in human skin in vivo studied by sensitive immunohistochemical methods. Carcinogenesis. 1983;4(6):745–750. doi: 10.1093/carcin/4.6.745. [DOI] [PubMed] [Google Scholar]
  6. Forster A. C., McInnes J. L., Skingle D. C., Symons R. H. Non-radioactive hybridization probes prepared by the chemical labelling of DNA and RNA with a novel reagent, photobiotin. Nucleic Acids Res. 1985 Feb 11;13(3):745–761. doi: 10.1093/nar/13.3.745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Franklin W. A., Lo K. M., Haseltine W. A. Alkaline lability of fluorescent photoproducts produced in ultraviolet light-irradiated DNA. J Biol Chem. 1982 Nov 25;257(22):13535–13543. [PubMed] [Google Scholar]
  8. Hayes F. N., Williams D. L., Ratlift R. L., Varghese A. J., Rupert C. S. Effect of a single thymine photodimer on the oligodeoxythymidylate-polydeoxyadenylate interaction. J Am Chem Soc. 1971 Sep;93(19):4940–4942. doi: 10.1021/ja00748a065. [DOI] [PubMed] [Google Scholar]
  9. Langer P. R., Waldrop A. A., Ward D. C. Enzymatic synthesis of biotin-labeled polynucleotides: novel nucleic acid affinity probes. Proc Natl Acad Sci U S A. 1981 Nov;78(11):6633–6637. doi: 10.1073/pnas.78.11.6633. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Leary J. J., Brigati D. J., Ward D. C. Rapid and sensitive colorimetric method for visualizing biotin-labeled DNA probes hybridized to DNA or RNA immobilized on nitrocellulose: Bio-blots. Proc Natl Acad Sci U S A. 1983 Jul;80(13):4045–4049. doi: 10.1073/pnas.80.13.4045. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Levine L., Seaman E., Hammerschlag E., Van Vunakis H. Antibodies to photoproducts of deoxyribonucleic acids irradiated with ultraviolet light. Science. 1966 Sep 30;153(3744):1666–1667. doi: 10.1126/science.153.3744.1666. [DOI] [PubMed] [Google Scholar]
  12. Mitchell D. L., Clarkson J. M. Use of synthetic polynucleotides to characterise an antiserum made against UV-irradiated DNA. Photochem Photobiol. 1984 Dec;40(6):743–748. doi: 10.1111/j.1751-1097.1984.tb04646.x. [DOI] [PubMed] [Google Scholar]
  13. Tan E. M. Antibodies to deoxyribonucleic acid irradiated with ultraviolet light: detection by precipitins and immunofluorescence. Science. 1968 Sep 27;161(3848):1353–1354. doi: 10.1126/science.161.3848.1353. [DOI] [PubMed] [Google Scholar]
  14. Tchen P., Fuchs R. P., Sage E., Leng M. Chemically modified nucleic acids as immunodetectable probes in hybridization experiments. Proc Natl Acad Sci U S A. 1984 Jun;81(11):3466–3470. doi: 10.1073/pnas.81.11.3466. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES