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DNA probes are fragments of DNA labeled with a reporter tag to enable their detection or purification. The resulting labeled DNA probes can then hybridize to target nucleic acid sequences through complementary base-pairing, and may be used to recover or identify these regions.
Radioisotopes, fluorophores, or small molecule binding partners like biotin or digoxigenin, are the most widely used reporter tags for labeling DNA probes. These labels can be attached to the probe DNA molecule via...
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Lanthanide-Complex-Enhanced Bioorthogonal Branched DNA Amplification.

Fang Zhao1, Yunpeng Guan2, Fei Su2

  • 1Department of Biomedical Engineering, Southern University of Science and Technology, Shenzhen 518055, China.

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|January 12, 2024
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Lanthanide-complex-enhanced bioorthogonal-branched DNA amplification (LEBODA) offers highly sensitive in situ nucleic acid detection in single cells. This novel method significantly improves signal amplification for detecting rare nucleic acid targets.

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Background:

  • Fluorescence in situ hybridization (FISH) is crucial for intracellular nucleic acid detection.
  • Low fluorescence intensity and background autofluorescence limit FISH sensitivity for low-copy nucleic acids.

Purpose of the Study:

  • To develop a highly sensitive method for in situ nucleic acid detection in single cells.
  • To overcome the limitations of traditional FISH techniques.

Main Methods:

  • Introduced lanthanide-complex-enhanced bioorthogonal-branched DNA amplification (LEBODA).
  • Employed click chemistry for signal amplification via bridge probes.
  • Incorporated high-density lanthanide complexes for secondary amplification.

Main Results:

  • LEBODA achieved a 4x signal enhancement compared to traditional probes for RNA detection.
  • Demonstrated superior detection of rare SARS-CoV-2-infected cells (as low as 20% infectious rate).

Conclusions:

  • LEBODA provides high sensitivity for in situ nucleic acid detection in single cells.
  • The adaptable LEBODA approach shows promise for DNA-FISH, single-molecule RNA-FISH, and other hybridization methods.
  • LEBODA offers broad potential for sensitive biomolecule detection.