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Related Experiment Videos

Membrane structural domains. Resolution limits using diphenylhexatriene fluorescence decay.

D A Barrow, B R Lentz

    Biophysical Journal
    |August 1, 1985
    PubMed
    Summary
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    Measuring fluorescence decay times of 1,6-diphenyl-1,3,5-hexatriene (DPH) in membranes can reveal lipid bilayer structures. However, resolving co-existing gel and fluid phases requires significant differences in DPH decay times.

    Area of Science:

    • Biophysics
    • Membrane Biophysics
    • Lipid Bilayer Dynamics

    Background:

    • 1,6-diphenyl-1,3,5-hexatriene (DPH) fluorescence decay can probe lipid bilayer structure.
    • Investigating structural domains requires precise measurement of multiple fluorescence decay times.

    Purpose of the Study:

    • To assess the feasibility of using DPH fluorescence decay to identify lipid bilayer structural domains.
    • To determine the precision limits for resolving multiple decay times in membrane suspensions.
    • To establish the minimum ratio of decay times needed for reliable resolution.

    Main Methods:

    • Reduced experimental errors in membrane suspension measurements (self-quenching, background fluorescence, turbidity).
    • Used phase and modulation techniques for fluorescence decay measurements.

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  • Correlated DPH fluorescence decay with lipid phase behavior in model systems.

    • Main Results:

    • Theoretical calculations showed decay time ratios must exceed 1.3 for reliable resolution.
    • Experimental validation demonstrated resolution of DPH decay from quinine sulfate when lifetime ratios were sufficient.
    • Mixtures of vesicles with distinct DPH decay times were resolved when lifetimes differed by >30%.
    • A short DPH fluorescence lifetime component was associated with gel-phase lipid vesicles.

    Conclusions:

    • DPH fluorescence lifetime heterogeneity can resolve co-existing gel and fluid phases in specific cases.
    • The precision of current methods limits the ability to detect subtle differences in lipid phases.
    • Further methodological improvements may enhance the utility of DPH fluorescence for membrane domain analysis.