Biology subjects

Heil, H. S.

Publications and source records attributed to Heil, H. S..

5 recordsLinked to original sources

bioRxiv2025

SMOLM-LFM: ratiometric single molecule orientation without polarizers

Single-molecule orientation and localization microscopy (SMOLM) enables the determination of molecular orientation, wobbling, and position. However, most SMOLM implementations rely on complex point spread function (PSF) fitting, which limits analysis throughput and introduces high computational cost. A way to overcome these limitations is to simplify the analysis using a ratiometric intensity estimation, often relying on polarization projections. While effective in 2D, extending these methods to 3D remains challenging. Here, we introduce a new ratiometric strategy for SMOLM in 3D. Building on the principles of Single Molecule Light Field Microscopy, which captures the 3D position information from a single snapshot by segmenting the back focal plane, we extend this strategy to orientation retrieval. Our approach uses the generalized 3D Stokes formalism to linearly decompose the intensity measurements across the light-field channels, allowing computationally-efficient estimations, while avoiding both complex PSF fitting and polarization projections. This framework, called SMOLM-LFM, enables 6D estimation of single molecules (3D position + 3D orientation) with a simplified optical setup and a large depth-of-field. We present the theoretical foundations, experimental implementation, and validation through measurements on calibration beads, single fluorophores, and cells, thereby demonstrating the methods potential and practical limitations.

bioengineering↗

bioRxiv2025

Smart Microscopy: Current Implementations and a Roadmap for Interoperability

Smart microscopy is transforming life sciences by automating experimental imaging workflows and enabling real-time adaptation based on feedback from images and other data streams. This shift increases throughput, improves reproducibility, and expands the functional capabilities of microscopes. However, the current landscape is highly fragmented. Academic researchers often develop custom solutions for specific scientific needs, while industry offerings are typically proprietary and tied to specific hardware. This diversity, while fostering innovation, also creates major challenges in interoperability, reproducibility, and standardization, which slows progress and adaption. This article presents a collaborative effort between academic and industry leaders to survey the current state of smart microscopy, highlight representative implementations, and identify common technical and organizational barriers. We propose a framework for greater interoperability based on shared standards, modular software design, and community-driven development. Our goal is to support collaboration across the field and lay the groundwork for a more connected, reusable, and accessible smart microscopy ecosystem. We conclude with a call to action for researchers, hardware developers, and institutions to join in building an open, interoperable foundation that will unlock the full potential of smart microscopy in life science research.

cell biology↗

bioRxiv2022

High-fidelity 3D live-cell nanoscopy through data-driven enhanced super-resolution radial fluctuation

In recent years, the development of new image analysis approaches has highlighted the possibility of recovering superresolution information from short sequences of wide-field images. Our recently developed method, SRRF (Super-Resolution Radial Fluctuations), enables long-term live-cell imaging beyond the resolution limit without specialized hardware. Here, we present eSRRF (enhanced-SRRF), a significant improvement over our initial method, enhancing image fidelity to the underlying structure and resolution. Especially, eSRRF uses automated data-driven parameter optimization, including an estimation of the number of frames necessary for optimal reconstruction. We demonstrate the improved fidelity of the images reconstructed with eSRRF and highlight its versatility and ease of use over a wide range of microscopy techniques and biological systems. We also extend eSRRF to 3D super-resolution microscopy by combining it with multi-focus microscopy (MFM), obtaining volumetric super-resolution imaging of live cells with acquisition speed of ~1 volume/second.

biophysics↗

bioRxiv2021

Mapping densely distributed membrane receptors in blood platelets with expansion microscopy

Interrogating small platelets and their densely packed, highly abundant receptor landscape is key to understand platelet clotting, a process that can save lives when stopping blood loss after an injury, but also kill when causing heart attack, stroke or pulmonary embolism. The underlying key receptor distributions and interactions, in particular the relevance of integrin clustering, are not fully understood is because of highly abundant and densely distributed GPIIb/IIIa receptors. This makes receptor distributions difficult to assess even by super-resolution fluorescence microscopy. Here, we combine dual-color expansion and confocal microscopy with colocalization analysis to assess platelet receptor organization without the need of a super-resolution microscope. We show that 4x expansion is highly straight-forward for super-resolution microscopy of platelets, while 10x expansion provides higher precision at the price of increased efforts in sample preparation and imaging. Quantifying various receptor colocalization scenarios we demonstrate that expansion microscopy can pinpoint receptor distributions and interactions in resting and activated platelets being superior to conventional methods that fail in such dense 3D scenarios with highly abundant receptors. We reveal the presence of GPIIb/IIIa clusters in resting platelets, which are not affected by platelet activation indicating that they contribute to the rapid platelet response during platelet clotting. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=80 SRC="FIGDIR/small/431449v3_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@c4e38forg.highwire.dtl.DTLVardef@37c1adorg.highwire.dtl.DTLVardef@8c8597org.highwire.dtl.DTLVardef@196bd69_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2017

Boosting The Localization Precision Of dSTORM By Biocompatible Metal-Dielectric Coated Glass Coverslips

Super-resolution techniques such as direct Stochastic Optical Reconstruction Microscopy (dSTORM) have become versatile and well-established tools for biological imaging over the last century. Here, we theoretically and experimentally show that clever combination of different fluorescence modalities allows further improvements. We found that the interaction of fluorophores with plasmonic surfaces boost super-resolution performance in dSTORM approaches as it allows for tailoring the excitation and emission properties. The strength of the approach is that no further specialized microscope setup is required as the described enhancement solely rely on metal-dielectric coated glass coverslips that are straightforward to fabricate. Such biocompatible plasmonic nanolayers enhance the signal-to-noise ratio of dSTORM, and thus sharpens the localization precision by a factor of two.

biophysics↗