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Time-resolved fluorometry: a sensitive method to quantify DNA-hybrids.

A C Syvänen, P Tchen, M Ranki

    Nucleic Acids Research
    |January 24, 1986
    PubMed
    Summary
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    Time-resolved fluorometry using europium (Eu) labels enables sensitive detection of nucleic acid hybrids. This method achieved a 0.3 pg detection limit for adenovirus DNA, applicable in clinical diagnostics.

    Area of Science:

    • Biochemistry
    • Molecular Biology
    • Analytical Chemistry

    Background:

    • Lanthanides, like europium (Eu), exhibit fluorescence upon UV excitation, with delayed emission up to 1 ms.
    • Europium serves as a sensitive label in various biological assays.
    • Time-resolved fluorometry offers a sensitive detection method for biological molecules.

    Purpose of the Study:

    • To apply time-resolved fluorometry for the detection of nucleic acid hybrids.
    • To develop a sensitive method for detecting specific DNA sequences.
    • To evaluate the utility of Eu-labeled antibodies in nucleic acid detection assays.

    Main Methods:

    • DNA probes were tagged with haptens (fluorene or sulfone).
    • Hybridization of tagged probes to target DNA was followed by a two-step immunological assay.

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  • The second antibody in the assay was labeled with europium (Eu).
  • Time-resolved fluorometry was used for Eu-based signal detection.

    • Main Results:

    • The developed method demonstrated quantitative detection of nucleic acid hybrids.
    • A detection limit of 0.3 pg for target adenovirus genomic DNA was achieved.
    • The Eu label was successfully employed in sandwich hybridization assays.
    • The method was applied to detect adenovirus in nasopharyngeal mucus samples.

    Conclusions:

    • Time-resolved fluorometry with Eu labels provides a highly sensitive method for nucleic acid hybrid detection.
    • The assay is quantitative and suitable for detecting specific DNA targets, including viral pathogens.
    • This technique holds potential for clinical diagnostics, such as analyzing patient samples for infectious agents like adenovirus.