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Related Experiment Videos

Probing single molecules in single living cells.

T A Byassee1, W C Chan, S Nie

  • 1Department of Chemistry, Indiana University, Bloomington 47405, USA.

Analytical Chemistry
|December 2, 2000
PubMed
Summary
This summary is machine-generated.

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Single-molecule detection in living cells is now possible using confocal fluorescence microscopy. This technique allows tracking of fluorescently tagged molecules like transferrin, rhodamine 6G, and oligonucleotides within cellular compartments.

Area of Science:

  • Cellular and Molecular Biology
  • Biophysics
  • Microscopy Techniques

Background:

  • Intracellular background fluorescence in living cells is higher than in buffer solutions.
  • Background fluorescence is continuous and stable, not interfering with single-molecule detection.
  • Confocal fluorescence microscopy enables spatially resolved intracellular measurements.

Purpose of the Study:

  • To demonstrate single-molecule detection within living human cells.
  • To investigate the intracellular trafficking of various fluorescently tagged molecules.
  • To validate confocal microscopy for intracellular single-molecule analysis.

Main Methods:

  • Utilized confocal fluorescence microscopy for intracellular imaging.
  • Employed externally tagged probe molecules for detection.

Related Experiment Videos

  • Analyzed single-molecule photon bursts from fluorescent probes.
  • Targeted specific cellular locations like cytoplasm and nucleus.

    • Main Results:

    • Successfully detected single molecules of transferrin, rhodamine 6G, and oligonucleotides in living HeLa cells.
    • Observed receptor-mediated endocytosis of transferrin.
    • Tracked potential-driven entry of rhodamine 6G.
    • Monitored passive uptake of fluorescently tagged oligonucleotides.

    Conclusions:

    • Confocal fluorescence microscopy is effective for single-molecule detection in living cells.
    • Intracellular background fluorescence does not impede single-molecule measurements.
    • The study provides insights into the cellular uptake mechanisms of different molecular probes.