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Total internal reflection fluorescence microscopy or TIRF is an advanced microscopic technique used to visualize fluorophores in samples close to a solid surface with a higher refractive index, such as a glass coverslip. TIRF only allows fluorophores in proximity to the solid surface to be excited. When light from a medium with a lower refractive index (such as air) hits the glass coverslip at a critical angle, the light undergoes total internal reflection stead of passing through the glass.
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Updated: May 10, 2025

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Targeted Delivery Inside the Cells Directly Visualized with Förster Resonance Energy Transfer (FRET).

Igor D Zlotnikov1, Natalya G Belogurova1, Elena V Kudryashova1

  • 1Faculty of Chemistry, Lomonosov Moscow State University, Leninskie Gory, 1/3, 119991 Moscow, Russia.

Polymers
|April 28, 2025
PubMed
Summary

A new Förster resonance energy transfer (FRET) assay effectively measures targeted drug delivery using polymeric micelles. This method visualizes drug penetration into bacterial cells, optimizing delivery systems for enhanced antibacterial activity.

Keywords:
FRETRFPbacteria cells targetingcurcuminpolymeric micellesumbelliferones

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Area of Science:

  • Biotechnology
  • Nanotechnology
  • Pharmacology

Background:

  • Targeted drug delivery systems are crucial for improving therapeutic efficacy and reducing side effects.
  • Polymeric micelles offer a promising platform for encapsulating and delivering therapeutic agents.
  • Developing efficient methods to evaluate drug delivery is essential for pharmaceutical research and development.

Purpose of the Study:

  • To establish a real-time Förster resonance energy transfer (FRET) based assay for evaluating targeted drug delivery using polymeric micelles.
  • To quantify drug delivery efficiency by measuring FRET efficiency between encapsulated drugs and a reporter protein.
  • To assess the impact of micellar composition on drug loading, delivery efficacy, and antibacterial activity.

Main Methods:

  • Utilized red fluorescent protein (RFP)-expressing *E. coli* as a test system.
  • Encapsulated fluorescent drugs (curcumin, umbelliferones) within polymeric micelles (heparin-based, chitosan-based).
  • Quantified drug delivery efficacy using FRET efficiency and visualized bacterial fluorescence on Petri dishes.

Main Results:

  • Polymeric micelles significantly enhanced the delivery and FRET efficiency of encapsulated drugs compared to free drugs.
  • Micellar composition influenced drug-polymer interactions, with heparin-based micelles showing stronger affinity for MUmb.
  • Curcumin-loaded micelles demonstrated substantial increases in RFP fluorescence, indicating effective delivery and interaction.
  • Micellar formulations exhibited enhanced antibacterial activity against *E. coli*, with Hep-OA micelles showing the most significant reduction in viability.
  • Synergistic effects were observed when combining micellar drugs with moxifloxacin.

Conclusions:

  • The developed FRET assay provides a versatile and accurate platform for evaluating targeted drug delivery systems.
  • Polymeric micelle composition critically affects drug loading, delivery efficiency, and therapeutic outcomes.
  • This approach facilitates the optimization of drug delivery systems for enhanced antibacterial efficacy and potential applications in treating intracellular infections.