Biology subjects

Balzarotti, F.

Publications and source records attributed to Balzarotti, F..

4 recordsLinked to original sources

bioRxiv2026

Super-resolved imaging of mRNA ultrastructure in cells

Messenger RNA (mRNA) is a central polymer of gene expression, whose ultrastructure and defining regulatory rules remain unclear. To visualize the ultrastructure of mRNA, we here develop Combi-PAINT, a powerful and generalizable method for combinatorial super-resolved DNA-PAINT multiplexing, which we combine with high-efficiency RNA labelling and MINFLUX microscopy. This approach enables the nanometer-precision tracing of mRNA in three dimensions in cells. We use Combi-PAINT to visualize multiple distinct mRNA species inside the human cell nucleus and cytoplasm, revealing their quantitative ultrastructures and transcript-specific molecular patterns. By bridging sequence specificity with nanometer resolution, we provide a new lens for studying the cellular life of mRNA.

biophysics↗

bioRxiv2023

A Fast Interferometric Beam Shaper for Multi-Emitter 3D MINFLUX

Beams of light that feature an intensity zero are essential to a variety of optical microscopy methods. Super-resolution techniques like STED and RESOLFT, together with localization strategies like MINFLUX and MINSTED, rely on accurate and fast displacements of such beams and their zeros. Extending these methods to the third dimension requires axial deflection, which, in contrast to lateral deflection, remains technologically challenging on the microsecond scale. Here, we present a fast general-purpose beam-shaping polarization interferometer that, instead of displacing the entire beam, enables such axial deflections by deforming the beam shape to deflect its zero. Based on this approach, we showcase a four-channel dual-color excitation system for three-dimensional MINFLUX imaging and tracking. We include first demonstrations of improved MINFLUX localization schemes that utilize the combination of distinct beam shapes and three-dimensional multi-emitter tracking. We believe that the presented approach will facilitate the broader adoption of three dimensional MINFLUX and provides a versatile basis for future implementations of advanced single-molecule localization methods.

biophysics↗

bioRxiv2022

Assessment of 3D MINFLUX data for quantitative structural biology in cells revisited

Prakash and Curd provide a re-analysis1 of individual datasets taken from our report2 demonstrating MINFLUX 3D imaging in cells. Their evaluation confirms the unique localization precision provided by MINFLUX2,3 featuring a standard deviation of {sigma} = 1-3 nm. We appreciate their confirmation and also welcome the opportunity to clarify their remaining points. The hitherto almost unconceivable 3D localization precision attained by MINFLUX is likely to hold the key to an all-optical dynamical structural biology.

biophysics↗

bioRxiv2019

MINFLUX nanoscopy delivers multicolor nanometer 3D-resolution in (living) cells

The ultimate goal of biological superresolution fluorescence microscopy is to provide three-dimensional resolution at the size scale of a fluorescent marker. Here, we show that, by localizing individual switchable fluorophores with a probing doughnut-shaped excitation beam, MINFLUX nanoscopy provides 1-3 nanometer resolution in fixed and living cells. This progress has been facilitated by approaching each fluorophore iteratively with the probing doughnut minimum, making the resolution essentially uniform and isotropic over scalable fields of view. MINFLUX imaging of nuclear pore complexes of a mammalian cell shows that this true nanometer scale resolution is obtained in three dimensions and in two color channels. Relying on fewer detected photons than popular camera-based localization, MINFLUX nanoscopy is poised to open a new chapter in the imaging of protein complexes and distributions in fixed and living cells.

biophysics↗