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Related Concept Videos

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Different fluorescence-based techniques are used to study the protein dynamics in living cells. These techniques include FRAP, FRET, and PET.
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Updated: Nov 6, 2025

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Optically Modulated and Optically Activated Delayed Fluorescent Proteins through Dark State Engineering.

Baijie Peng1, Ryan Dikdan1, Shannon E Hill1

  • 1School of Chemistry & Biochemistry and Petit Institute for Biosciences and Bioengineering, Georgia Institute of Technology, Atlanta, Georgia 30332-0400, United States.

The Journal of Physical Chemistry. B
|May 12, 2021
PubMed
Summary
This summary is machine-generated.

Researchers engineered yellow fluorescent proteins to create new emitters with unique delayed fluorescence. These optically modulated and activated proteins enable highly sensitive, background-free biological imaging and detection.

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Area of Science:

  • Biophysics
  • Molecular Biology
  • Optical Engineering

Background:

  • Modulating fluorescent protein emission enhances sensitivity in biological imaging.
  • Yellow fluorescent proteins (YFPs) possess dark states that can be modulated by secondary illumination.

Purpose of the Study:

  • To engineer yellow fluorescent proteins (YFPs) with distinct modulation profiles and optically gated, delayed fluorescence.
  • To develop novel fluorescent proteins for background- and reference-free, high-sensitivity cellular imaging.

Main Methods:

  • Engineering EYFP and mVenus YFPs through single point mutations and double mutations.
  • Utilizing secondary illumination (near-IR coexcitation) to induce optically activated delayed fluorescence (OADF).
  • Spectroscopic analysis to characterize modulation pathways and dark states.

Main Results:

  • Identified single point mutations creating longer-lived modulatable dark states and double mutations rendering EYFP unmodulatable.
  • Demonstrated OADF via secondary laser-induced reverse intersystem crossing (RISC) from a triplet state.
  • Developed Optically Modulated and Optically Activated Delayed Fluorescent Proteins (OMFPs and OADFPs) with unique modulation spectra and OADF properties.

Conclusions:

  • Engineered YFPs offer distinct modulation profiles and optically gated delayed fluorescence for advanced bioimaging.
  • OMFPs and OADFPs provide a pathway for background-free, high-sensitivity detection.
  • Structural and spectroscopic data offer mechanistic insights for designing future optically modulated fluorescent proteins.