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Related Experiment Videos

Flow cytometry-based biosensor for detection of multivalent proteins.

X Song1, J Shi, B Swanson

  • 1Bioscience Division, Los Alamos National Laboratory, Los Alamos, New Mexico 87544, USA. xsong@lanl.gov

Analytical Biochemistry
|August 10, 2000
PubMed
Summary
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A new biosensor detects cholera toxin (CT) using microsphere-based flow cytometry and fluorescence resonant energy transfer (FRET). This sensitive method identifies low picomolar concentrations of CT quickly, aiding in toxin detection and kinetic studies.

Area of Science:

  • Biomedical Engineering
  • Analytical Chemistry
  • Biophysics

Background:

  • Cholera toxin (CT) is a critical virulence factor of Vibrio cholerae.
  • Sensitive and rapid detection of CT is crucial for diagnosing cholera and monitoring outbreaks.
  • Existing detection methods may lack sensitivity, speed, or convenience.

Purpose of the Study:

  • To develop a novel microsphere-based flow cytometric assay for sensitive cholera toxin detection.
  • To utilize fluorescence resonant energy transfer (FRET) for signal generation.
  • To enable rapid kinetic studies of CT interactions.

Main Methods:

  • Fabrication of supported phospholipid bilayers with ganglioside GM1 on microspheres.
  • Labeling GM1 with both fluorescence donor and acceptor fluorophores.

Related Experiment Videos

  • Detection of CT using a commercial flow cytometer measuring FRET-induced fluorescence changes.
  • Optimization of GM1 surface density and fluorophore ratios.

    • Main Results:

    • Developed a flow cytometry-based biosensor for CT detection.
    • Achieved sensitive detection of CT at concentrations below 10 pM within 30 minutes.
    • Demonstrated simultaneous double-fluorescence changes indicative of multivalent CT interactions.
    • Optimized surface parameters for enhanced assay sensitivity.

    Conclusions:

    • The developed microsphere-based flow cytometry assay offers a convenient and sensitive method for CT detection.
    • The FRET-based approach allows for rapid kinetic analysis of multivalent binding interactions.
    • This technology holds promise for point-of-care diagnostics and epidemiological surveillance of cholera.