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Labeling DNA Probes

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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Genetically-encoded Molecular Probes to Study G Protein-coupled Receptors
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A cell-based computational modeling approach for developing site-directed molecular probes.

Jing-Yu Yu1, Nan Zheng, Gerta Mane

  • 1Department of Pharmaceutical Sciences, University of Michigan College of Pharmacy, Ann Arbor, Michigan, United States of America.

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|March 3, 2012
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Summary

This study developed a computational and experimental method to predict where small molecules accumulate in lung cells. Mitochondrial accumulation of specific cations enhances retention in upper airway cells after intratracheal administration.

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Area of Science:

  • Pharmacology
  • Biophysics
  • Computational Biology

Background:

  • Predicting small molecule absorption and retention in cells is crucial for developing targeted bioimaging and therapeutic agents.
  • Site-directed chemical agents require understanding local cellular uptake and retention patterns.

Purpose of the Study:

  • To develop and validate an integrative approach combining computational modeling, in vitro assays, and in vivo studies to predict local absorption and retention of small molecules in lung epithelial cells.
  • To optimize the targeting of small molecule probes to the upper airway epithelial cells.

Main Methods:

  • Utilized a multiscale computational model of the lung for in silico screening of small molecules.
  • Employed in vitro cell-based assays, including differentiated cell monolayers and air-liquid interface cultures of primary human bronchial epithelial cells, to assess local absorption and intracellular retention.
  • Conducted in vivo biodistribution studies with bioimaging probes to validate computational models and in vitro findings after local and systemic administration.

Main Results:

  • In silico simulations accurately predicted in vivo experimental outcomes.
  • Mitochondrial accumulation of membrane-permeant, hydrophilic cations was identified as a key factor for maximizing local exposure and retention.
  • Demonstrated that this approach can effectively target the upper airways following intratracheal administration.

Conclusions:

  • An integrative computational and experimental strategy can effectively model and predict local small molecule absorption and retention in cellular contexts, specifically the lung.
  • Mitochondrial accumulation of specific cation types offers a viable strategy for enhancing local retention of chemical agents in the upper airways.