title: Terbium fluorescence as a sensitive, inexpensive probe for UV-induced damage in nucleic acids.
abstract: Much effort has been focused on developing methods for detecting damaged nucleic acids. However, almost all of the proposed methods consist of multi-step procedures, are limited, require expensive instruments, or suffer from a high level of interferences. In this paper, we present a novel simple, inexpensive, mix-and-read assay that is generally applicable to nucleic acid damage and uses the enhanced luminescence due to energy transfer from nucleic acids to terbium(III) (Tb(3+)). Single-stranded oligonucleotides greatly enhance the Tb(3+) emission, but duplex DNA does not. With the use of a DNA hairpin probe complementary to the oligonucleotide of interest, the Tb(3+)/hairpin probe is applied to detect ultraviolet (UV)-induced DNA damage. The hairpin probe hybridizes only with the undamaged DNA. However, the damaged DNA remains single-stranded and enhances the intrinsic fluorescence of Tb(3+), producing a detectable signal directly proportional to the amount of DNA damage. This allows the Tb(3+)/hairpin probe to be used for sensitive quantification of UV-induced DNA damage. The Tb(3+)/hairpin probe showed superior selectivity to DNA damage compared to conventional molecular beacons probes (MBs) and its sensitivity is more than 2.5 times higher than MBs with a limit of detection of 4.36±1.2 nM. In addition, this probe is easier to synthesize and more than eight times cheaper than MBs, which makes its use recommended for high-throughput, quantitative analysis of DNA damage.
authors.affiliation: Department of Chemistry, University of Alberta, Edmonton, AB, Canada.
authors.forename: Amira F
authors.initials: AF
authors.lastname: El-Yazbi
authors.forename: Glen R
authors.initials: GR
authors.lastname: Loppnow
mesh_terms: D004249:DNA Damage / Q000528:radiation effects*; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000737:chemistry* / Q000191:economics / Q000592:standards; D009696:Nucleic Acids / Q000032:analysis / Q000528:radiation effects; D009841:Oligonucleotides / Q000032:analysis* / Q000528:radiation effects*; D013725:Terbium / Q000737:chemistry* / Q000191:economics / Q000592:standards; D014466:Ultraviolet Rays / Q000009:adverse effects*
chemical_list: D005456:Fluorescent Dyes; D009696:Nucleic Acids; D009841:Oligonucleotides; D013725:Terbium