Novel ‘smart’ molecules promise better and cheaper microscopy images

04 Mar 2026

Chalmers-developed chemicals achieve higher resolution than 4-photon microscopy, even using 2-photon method.

High-resolution images without expensive microscopy equipment. The aim of the EU-funded project “4 for 2” was to develop a new category of smart molecules for labelling cells, enabling more efficient use of light from simpler microscopes. After three years, it is time to review the results.

Multiphoton microscopy is used in biomedical research to study cells and tissues. Today, so-called two-photon microscopy is used to study processes within cells, but the technique has limitations in terms of image resolution. Four-photon microscopy provides images with higher resolution. However, such instruments are very expensive and, when studying biological material, the powerful laser light required can damage samples.

“In this project, we have developed molecules to visualise molecular-level details and monitoring processes using the more common two-photon microscopy technique. These molecules have the capacity to achieve higher resolution than with four-photon microscopy, although two-photon microscopy is used,” said project coordinator Prof. Joakim Andréasson, from the Department of Chemistry & Chemical Engineering at Sweden’s Chalmers University of Technology.

Andréasson added, “In the long term, results from studies of this kind may provide new insights into diseases, pharmaceuticals and the very smallest building blocks of life.”

Resolution doubled

The researchers synthesized the molecules by linking fluorescent dyes with so-called molecular photoswitches, small molecules that change their properties when illuminated. The idea was to demonstrate that this combination could be used to achieve improved resolution.

When cell samples stained with these molecules are examined using two-photon microscopy, the molecules behave as if they were illuminated with the light used in four-photon microscopy, but achieving twice as good resolution compared to the latter teqchnique.

“In a recently published study in Nature Communications, we showed that the molecules exhibit four-photon behaviour, even when using two-photon microscopy. Further studies are required to optimise the technique, particularly for investigations in biologically relevant environments, such as live cells,” said Andréasson.

The “4 for 2” project was awarded €2.3 million over three years by the EU Pathfinder Open initiative. The project is a collaboration between four universities: Chalmers University of Technology, the University of Gothenburg, KU Leuven in Belgium and the University of Huelva in Spain. Joakim Andréasson describes the projects broad, combined expertise ? ranging from specialists in organic chemistry, spectroscopy and photo physics to chemical biology ?as its key strength and a major factor in its success.

“The substantial project funding also gave us the opportunity to focus on ensuring high quality results without time pressure. Now that the project is drawing to a close, we can see that we have been very successful in delivering on what we promised in our grant proposal, while also generating additional results for future studies,” said Andréasson.