Having a microscope that can magnify and enhance the tiniest details of biological structures can reveal a world beyond the limits of conventional resolution. This is precisely what enhanced super-resolution radial fluctuations (eSRRF) bring to the field of microscopy.
What is SRRF?
In the field of microscopy, two powerful techniques, light, and electron microscopy, provide complementary information on biological samples across broad size regimes. However, a gap between light and electron microscopy prompts the physicists to search for ways to overcome the resolution barrier.
Over the last two decades, super-resolution microscopy (SRM) developments have allowed the unprecedented observation of nanoscale structures in biological systems. SRM refers to a series of fluorescence imaging techniques in optical microscopy that enables images to have higher resolutions than those imposed by the diffraction limit.
Different SRM methods have evolved and are being widely applied in biomedical research. Some techniques can have nanometer-scale molecular resolution, while others are geared towards volumetric 3D multi-color or fast live-cell imaging.
In particular, super-resolution radial fluctuation (SRRF) is a versatile technique that achieves live-cell SRM on a wide range of microscopy platforms. As a computational approach to SRM, SRRF is highly accessible to life science researchers since it allows using common microscopes. Users are enabled to generate super-resolution images using the same equipment and methods they routinely employ for their studies without preparing specialized samples or reagents.
Since its inception, various adaptations of SRRF have been proposed, like those based on a combination with other advanced imaging techniques or on introducing additional data preprocessing steps. It is currently a widely used high-density reconstruction algorithm, as proven by a significant uptake by the scientific community.
However, obtaining 3D SRM in live-cell microscopy is still challenging for experts. Since the current implementations of 3D super-resolution methods come at the expense of long acquisition times and limited axial range, major technical expertise is usually required.
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Turning the Invisible Visible
At Instituto Gulbenkian de Ciência (IGC), a team of scientists presents a new implementation of the SRRF approach and highlights its improved capabilities. Termed eSRRF, the new technique redefines some of the original fundamental principles of SRRF to achieve improved image quality in the reconstructions.
The eSRRF is built upon the success of the SRRF method. With automated data-driven parameter optimization, the eSRRF determines the optimal number of frames required for reconstruction, providing a hassle-free and efficient imaging experience.
Additionally, eSRRF transcends dimensions by teaming up with multi-focus microscopy, leading to 3D super-resolution. Because of this, volumetric snapshots of live cells are captured at a breathtaking speed of around one volume per second. Regarding research openness and usability, eSRRF was designed to be user-friendly and seamlessly integrated with different microscopy techniques and biological systems.
According to research first author Hannah Heil, eSRRF is not just about enhancing image resolution. Instead, it empowers researchers to optimize the results based on quantitative image quality measures. In other words, the new method provides researchers with dynamic tools that can adapt to their needs.
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