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Total Internal Reflection Fluorescence Microscopy

Total Internal Reflection Fluorescence Microscopy

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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Related Experiment Video

Updated: Feb 17, 2026

Live Images of GLUT4 Protein Trafficking in Mouse Primary Hypothalamic Neurons Using Deconvolution Microscopy

Live Images of GLUT4 Protein Trafficking in Mouse Primary Hypothalamic Neurons Using Deconvolution Microscopy

Published on: December 7, 2017

Total Internal Reflection Fluorescence Microscopy to Study GLUT4 Trafficking.

Sebastian Wasserstrom¹, Björn Morén¹, Karin G Stenkula²

¹Department of Experimental Medical Science, Lund University, BMC C11, Lund, 22 184, Sweden.

Methods in Molecular Biology (Clifton, N.J.)

|December 9, 2017

Summary

Total internal reflection fluorescence (TIRF) microscopy visualizes real-time plasma membrane events. This method is crucial for studying glucose transporter type 4 (GLUT4) translocation in adipocytes, providing detailed insights into cellular processes.

Keywords:

Fluorescence GLUT4 GLUT4 storage vesicles GSV Microscopy Primary adipocytes TIRF

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High-resolution Spatiotemporal Analysis of Receptor Dynamics by Single-molecule Fluorescence Microscopy

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Related Experiment Videos

Last Updated: Feb 17, 2026

Live Images of GLUT4 Protein Trafficking in Mouse Primary Hypothalamic Neurons Using Deconvolution Microscopy

Live Images of GLUT4 Protein Trafficking in Mouse Primary Hypothalamic Neurons Using Deconvolution Microscopy

Published on: December 7, 2017

Quantitative Measurement of GLUT4 Translocation to the Plasma Membrane by Flow Cytometry

Quantitative Measurement of GLUT4 Translocation to the Plasma Membrane by Flow Cytometry

Published on: November 7, 2010

High-resolution Spatiotemporal Analysis of Receptor Dynamics by Single-molecule Fluorescence Microscopy

High-resolution Spatiotemporal Analysis of Receptor Dynamics by Single-molecule Fluorescence Microscopy

Published on: July 25, 2014

Area of Science:

Cell biology
Biophysics
Physiology

Background:

Total internal reflection fluorescence (TIRF) microscopy enables real-time observation of cellular events near the plasma membrane.
GLUT4 (glucose transporter type 4) translocation is a critical process in adipocytes for regulating blood glucose levels.
Previous studies have utilized TIRF microscopy to investigate GLUT4 trafficking.

Purpose of the Study:

To detail a standardized procedure for studying GLUT4 trafficking in isolated primary adipocytes using TIRF microscopy.
To provide a methodological framework for researchers investigating glucose transport dynamics.

Main Methods:

Isolation of primary adipocytes.
Application of Total internal reflection fluorescence (TIRF) microscopy.
Real-time monitoring of GLUT4 vesicle movement and fusion events at the plasma membrane.

Main Results:

Demonstration of successful real-time visualization of GLUT4 translocation events.
Quantitative analysis of GLUT4 vesicle dynamics, including fusion frequency and residence time.
Validation of the TIRF microscopy protocol for studying GLUT4 trafficking in primary adipocytes.

Conclusions:

TIRF microscopy is an effective technique for studying GLUT4 trafficking in adipocytes.
The described protocol facilitates detailed investigation of glucose transporter dynamics at the cellular level.
This methodology can advance understanding of metabolic diseases related to insulin resistance.