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Trinity College Dublin lights up cellular activity

20 Feb 2024

Fluorescence lifetime imaging with new dye visualises multiple biological environments.

A project at Trinity College Dublin has developed a novel route to fluorescence imaging of multiple distinct biological environments.

Published in Chem, the breakthrough could be of immediate benefit in medical applications, with other uses likely to follow in chemistry and biology.

The work forms part of the ongoing effort to enhance to versatility and accuracy of fluorescence imaging techniques, work which has included advances in both optics and chemistry.

Recent examples include work at MIT to detect fluorescence in deep locations within tissues, through the use of three distinct laser wavelengths and an oscillating excitation pattern.

Wavelength engineering is also the approach of a project at Washington University in St. Louis employing dual excitation fluorescence to capitalize on the wavelength-dependent attenuation of light in tissue, and exploit it as a means of determining fluorophore depth.

The Trinity College Dublin study has now developed fluorescent colour-changing dyes that, for the first time, can be used to simultaneously visualise multiple distinct biological environments using only one singular dye.

"Bioimaging relies on 'on/off' dyes, where the dyes only emit light under one set of conditions but are otherwise switched off," commented Adam Henwood from the college's Trinity Biomedical Sciences Institute (TBSI).

"This is extremely useful, but it does mean that you can only look at one place at a time under your microscope. The exciting part about this work is that our dyes hit a sweet spot that gives them distinctive on/off/on properties, and crucially we can both observe and differentiate these different 'on' states.

New understanding of cell metabolism

Accurately timing these distinct behaviors using fluorescence lifetime imaging (FLIM) can yield more valuable information about the tissues being imaged.

The dyes are encapsulated within delivery vessels, and light from those vessels takes marginally longer to reach the observing microscope than light from within the cells. By collecting enough light signals, this information can rapidly build up precise 3D images of the two different dye environments. The time differences are small, just a few billionths of a second according to the Trinity team, but the new method is sensitive enough to capture it.

To test the approach the project used its dyes to image cellular lipid droplets, one example of the organelles that make up living cells in most complex organisms. FLIM was able to differentiate two distinct fluorescence emissions from human breast cancer cells after uptake of the dye, from extracellular and intracellular species.

This proof of concept opens up further potential applications, including use of the dyes as sensors for hazardous environmental pollutants or using their controlled emission to power further chemical reactions, as well as improving the understanding of cell metabolism.

"The closer we look at the molecule-cell interface, and the better we can see in real time how molecules diffuse from place to place inside the cell nanomachinery, the closer we get to realising Richard Feynman's dream of understanding everything that living things do from the wiggling and jiggling of atoms," commented Damien Thompson from Science Foundation Ireland's Research Centre for Pharmaceuticals (SSPC).

"But only recently have researchers had sufficient experimental and computational resources to track these motions and vibrations in complex biological environments. This exciting new work demonstrates more specific, high contrast imaging of subcellular dynamics, which will in turn enable researchers to develop more effective drug formulations with reduced side effects."

LaCroix Precision OpticsCHROMA TECHNOLOGY CORP.LASEROPTIK GmbHHÜBNER PhotonicsMad City Labs, Inc.AlluxaTRIOPTICS GmbH
© 2024 SPIE Europe
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