Biology subjects

van Tartwijk, F. W.

Publications and source records attributed to van Tartwijk, F. W..

4 recordsLinked to original sources

bioRxiv2025

SIMple: A fibre-based platform for accessible structured illumination microscopy

Structured illumination microscopy can be used to achieve optical sectioning and super resolution in fluorescence images, reducing out-of-focus light and increasing the resolution beyond the diffraction limit, without the need for specialised detection optics. However, the complex illumination path is difficult to build and align. We present an illumination path based on fibre-optic components for both splitting and phase-shifting the illumination light. This enables a SIMple and compact "Plug&Play" modality which substantially reduces the time and alignment required when adding the optics to an existing widefield instrument. The system is capable of optical sectioning imaging at camera-limited frame-rates using multiple excitation wavelengths simultaneously, as demonstrated by imaging fixed and live biological samples at 561 and 491 nm. Super-resolution imaging of fixed samples on a very compact, self-contained microscope is also demonstrated: illumination is coupled in by fibre to a lightweight frame with dimensions of just 300 x 450 x 300 mm3, enabling easy transportation and use in laboratories with limited space. Characterisation of the system using bead analysis shows a resolution of 168 and 172 nm at 491 and 561 nm, respectively, an improvement by a factor of 1.91 and 1.92 compared to widefield, with a field of view of 100 x 100 {micro}m2.

bioengineering↗

bioRxiv2025

An Enhanced Mountain Climbing Search Algorithm to Enable Fast and Accurate Autofocusing in High Resolution Fluorescence Microscopy

Accurate and efficient autofocusing is essential for the automation of fluorescence microscopy, but background noise and shallow depth of field at high magnifications make autofocusing particularly challenging. Here, we present a fast and accurate autofocus algorithm to address these challenges. It is highly effective for high-magnification imaging, while performing equally well for low-magnification imaging tasks. The method is based on the mountain climbing search algorithm and yields improvements on autofocusing precision of up to 200-fold over current methods, whilst offering competitive speed and greatly extended search ranges. Our approach is broadly applicable: it demonstrated good stability and reproducibility across magnifications ranging from 20X to 100X, excels in both live cell imaging and high-resolution fixed sample imaging, and it is compatible with various microscopy techniques without the need for fiducial markers or hardware modifications on existing microscopes. To maximise its accessibility, we constructed a user-friendly interface compatible with the widely used Micromanager software. It generalises well across various imaging modalities and hardware platforms, making it particularly suitable for use in high-resolution screening of candidate drugs.

cell biology↗

bioRxiv2024

A high-resolution microscopy system for biological studies of cold-adapted species under physiological conditions

The fundamental processes governing life are sensitively dependent on temperature. Whilst much is known about the constraints on how proteins operate at 37{degrees}C, little knowledge exists about how biological function is maintained sub-zero temperature conditions, where proteins are less stable and oxidative damage is high. However, almost 90% of habitable environments on Earth are permanently below 5{degrees}C (i.e. the deep sea and polar regions). This means that we do not understand how a large and diverse proportion of the global biome functions. To address this question at the cellular level, tools are required for imaging biological systems at high resolution under physiological conditions. This poses severe technical challenges that cannot be addressed with traditional optical microscopy techniques. High-resolution imaging objectives require short working distances and the use of immersion media, which lead to rapid heat transfer from the microscope to the sample. This affects the viability of live specimens and the interpretability of the results when the sample function optimally at low temperatures. Condensation and temperature-induced shrinking of components pose further challenges, reducing image resolution and contrast. Here, we address these issues and provide a method for high-fidelity imaging of live biological samples at temperatures of around, or below, 0{degrees}C. Our method is compatible with different microscopy modalities, including super-resolution imaging. It relies on hardware additions to traditional microscopy systems that can be straightforwardly implemented, namely, a cooling collar, 10% ethanol as an immersion medium, and nitrogen flow to mitigate condensation. We demonstrate the method in live cell cultures derived from Antarctic fish species and highlight the need to maintain physiological conditions for these fragile biological samples. Future applications are diverse and include evolutionary biology and the study of cold-adapted organisms, as well as cellular biophysics and several applications in biotechnology.

biophysics↗

bioRxiv2022

Mutation of the ALS/FTD-associated RNA-binding protein FUS alters axonal cytoskeletal organisation

Aberrant condensation and localisation of the RNA-binding protein fused in sarcoma (FUS) occur in variants of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). ALS is also associated with cytoskeletal defects, genetically and through observations of compromised axonal transport. Here, we asked whether compromised axonal cytoskeletal organisation is an early feature of FUS-associated ALS/FTD. We used an ALS-associated mutant FUS(P525L) and the FTD-mimic hypomethylated FUS, FUS(16R), to investigate the common and distinct cytoskeletal changes found in these two reported Xenopus models. Combining a novel atomic force microscopy (AFM)-based approach for in vitro cytoskeletal characterisation and in vivo axonal branching analysis, we found that mutant FUS reduced actin density in the dynamically remodelling growth cone, and reduced axonal branch complexity. We furthermore found evidence of an axon looping defect for FUS(P525L). Therefore, we show that compromised actin remodelling is potentially an important early event in FUS-associated pathogenesis. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/510780v2_ufig1.gif" ALT="Figure 1"> View larger version (56K): org.highwire.dtl.DTLVardef@1e9a92aorg.highwire.dtl.DTLVardef@161076eorg.highwire.dtl.DTLVardef@12edf0forg.highwire.dtl.DTLVardef@365785_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗