Biology subjects

Lamb, J. R.

Publications and source records attributed to Lamb, J. R..

10 recordsLinked to original sources

bioRxiv2026

Index-agnostic oblique plane light sheet microscopy of centimetre-scale cleared tissues at subcellular resolution

We present cleared-tissue direct-view oblique plane microscopy (CtDvOPM), which enables optically sectioned subcellular resolution imaging of centimetre-scale tissues at high throughput over the full range of clearing media refractive indices (n = 1.33-1.56). CtDvOPM can image conventionally-mounted expanded, aqueous or non-aqueous cleared tissue samples at up to 2 {micro}m lateral by 14 {micro}m axial resolution over a 10.1 mm x 25.0 mm x 6.8 mm (x x y x z) sample volume without image tiling, at up to 400 million voxels per second.

biophysics↗

bioRxiv2025

Maximising imaging volumes of expanded tissues for inverted fluorescence microscopy

Expansion microscopy (ExM) has enabled nanoscale imaging of tissues by physically enlarging biological samples in a swellable hydrogel. However, the increased sample size and water-based environment pose challenges for deep imaging using conventional inverted confocal microscopes, particularly due to the limited working distance of high-numerical aperture (NA) water immersion objectives. Here, we introduce a practical imaging alternative that utilizes an inverted water-dipping objective and a refractive-index-matched optical path using fluorinated ethylene propylene (FEP) film. Through point spread function (PSF) measurements and simulations, we show that the FEP film introduces predominantly defocus-like wavefront profiles characteristic of high NA systems, which result in an easily correctable axial shift of the focal plane. To ensure stable immersion and refractive index continuity, we use an arrangement relying on an FEP film, Immersol W, water and a FEP-based imaging dish. This configuration achieves sub-micron lateral and axial resolution, supports large tile-scan acquisitions, and maintains image quality across depths exceeding 800 {micro}m. We validate the system by imaging 4x-expanded U2OS cells and human cerebral organoids. Our approach provides a low-cost, plug-and-play solution for high-resolution volumetric imaging of expanded samples using standard inverted microscopes.

biophysics↗

bioRxiv2025

A deconvolution solution to aliasing confusion in apparently undersampled oblique plane microscopy

In oblique plane microscopy, a remote refocussing system is used to create an aberration-free 3D image of the object, from which an inclined plane is sampled using a tilted imaging system. As a result, unlike in conventional microscopes, the point spread function (PSF) and optical transfer function (OTF) are tilted with respect to the optical axis of the imaging system. This suggests that a small sample-scanning step size is required to satisfy the Nyquist-Shannon sampling criterion. Recently, we have shown that a careful choice of apparent undersampling leads to aliased OTF copies that can be separated and stitched together in Fourier space to form a properly sampled OTF. In our demonstrations, this led to acquisition speed gains of between twoand four-fold. Here, we introduce a deconvolution-based method to reconstruct properly sampled volumes from apparently undersampled datasets that outperforms our previous Fourier-stitching approach.

biophysics↗

bioRxiv2025

Direct-view oblique plane microscopy

The ability to rapidly image mesoscopic samples is critically important for many areas of biological research. Owing to its high through-put and gentle, volumetric imaging nature, light-sheet fluorescence microscopy (LSFM) is an attractive modality to image such samples. However, the orthogonal dual-objective geometry of LSFM makes sample mounting challenging. Oblique plane microscopy (OPM) circumvents these issues by achieving light-sheet imaging through a single objective near the sample, with a further two objectives acting to image a tilted, oblique plane in the specimen. However, at low magnification, as required for mesoscopic imaging, conventional OPM systems suffer from a complicated and inefficient light path and/or a limited numerical aperture. Here we present a simple method for mesoscopic OPM that enables efficient light collection at any numerical aperture. By placing the camera directly in the remote space, the oblique plane can be imaged without the need for a third microscope system. Using a commercially available camera with 1.4 {micro}m pixels, we demonstrate imaging over a 5.3 x 3 x 2.6 mm3 field of view with a lateral resolution of [~]2 {micro}m and axial resolution of [~]22 {micro}m. Our technique is then used to image a 4x-expanded brain organoid.

biophysics↗

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↗

bioRxiv2024

Enzyme-Responsive DNA Condensates

Membrane-less compartments and organelles are widely acknowledged for their role in regulating cellular processes, and there is an urgent need to harness their full potential as both structural and functional elements of synthetic cells. Despite rapid progress, synthetically recapitulating the nonequilibrium, spatially distributed responses of natural membrane-less organelles remain elusive. Here we demonstrate that the activity of nucleic-acid cleaving enzymes can be localised within DNA-based membrane-less compartments by sequestering the respective DNA or RNA substrates. Reaction-diffusion processes lead to complex nonequilibrium patterns, dependent on enzyme concentration. By arresting similar dynamic patterns, we spatially organise different substrates in concentric sub-compartments, which can be then selectively addressed by different enzymes, demonstrating spatial distribution of enzymatic activity. Besides advancing our ability to engineer advanced biomimetic functions in synthetic membrane-less organelles, our results may facilitate the deployment of DNA-based condensates as microbioreactors or platforms for the detection and quantitation of enzymes and nucleic acids.

synthetic biology↗

bioRxiv2021

A genome-wide association study of serum proteins reveals shared loci with common diseases

With the growing number of genetic association studies, the genotype-phenotype atlas has become increasingly more complex, yet the functional consequences of most disease associated alleles is not understood. The measurement of protein level variation in solid tissues and biofluids integrated with genetic variants offers a path to deeper functional insights. Here we present a large-scale proteogenomic study in 5,368 individuals, revealing 4,113 independent associations between genetic variants and 2,099 serum proteins, of which 37% are previously unreported. The majority of both cis- and trans-acting genetic signals are unique for a single protein, although our results also highlight numerous highly pleiotropic genetic effects on protein levels and demonstrate that a proteins genetic association profile reflects certain characteristics of the protein, including its location in protein networks, tissue specificity and intolerance to loss of function mutations. Integrating protein measurements with deep phenotyping of the cohort, we observe substantial enrichment of phenotype associations for serum proteins regulated by established GWAS loci, and offer new insights into the interplay between genetics, serum protein levels and complex disease.

genomics↗

bioRxiv2021

The SARS-CoV-2 nucleocapsid protein associates with the replication organelles before viral assembly at the Golgi/ERGIC and lysosome-mediated egress

Despite being the target of extensive research efforts due to the COVID-19 pandemic, relatively little is known about the dynamics of SARS-CoV-2 replication within cells. We investigate and characterise the tightly orchestrated sequence of events during different stages of the infection cycle by visualising the spatiotemporal dynamics of the four structural proteins of SARS-CoV-2 at high resolution. The nucleoprotein is expressed first and accumulates around folded ER membranes in convoluted layers that connect to viral RNA replication foci. We find that of the three transmembrane proteins, the membrane protein appears at the Golgi apparatus/ERGIC before the spike and envelope proteins. Relocation of the lysosome marker LAMP1 towards the assembly compartment and its detection in transport vesicles of viral proteins confirm an important role of lysosomes in SARS-CoV-2 egress. These data provide new insights into the spatiotemporal regulation of SARS-CoV-2 assembly, and refine current understanding of SARS-CoV-2 replication.

microbiology↗

bioRxiv2020

Human serum proteome profoundly overlaps with genetic signatures of disease

Circulating proteins are prognostic for human outcomes including cancer, heart failure, brain trauma and brain amyloid plaque burden. A deep serum proteome survey recently revealed close associations of serum protein networks and common diseases. The present study reveals unprecedented number of individual serum proteins that overlap genetic signatures of diseases emanating from different tissues of the body. Here, 54,469 low-frequency and common exome-array variants were compared with 4782 protein measurements in the serum of 5343 individuals of the deeply annotated AGES Reykjavik cohort. Using a study-wide significant threshold, 2019 independent exome array variants affecting levels of 2135 serum proteins were identified. These variants overlapped genetic loci for hundreds of complex disease traits, emphasizing the emerging role for serum proteins as biomarkers of and potential causative agents of multiple diseases.

genetics↗

bioRxiv2019

Deep serum proteomics reveal biomarkers and causal candidates for type 2 diabetes

The prevalence of type 2 diabetes mellitus (T2DM) is expected to increase rapidly in the next decades, posing a major challenge to societies worldwide. The emerging era of precision medicine calls for the discovery of biomarkers of clinical value for prediction of disease onset, where causal biomarkers can furthermore provide actionable targets. Blood-based factors like serum proteins are in contact with every organ in the body to mediate global homeostasis and may thus directly regulate complex processes such as aging and the development of common chronic diseases. We applied a data-driven proteomics approach measuring serum levels of 4,137 proteins in 5,438 Icelanders to discover novel biomarkers for incident T2DM and describe the serum protein profile of prevalent T2DM. We identified 536 proteins associated with incident or prevalent T2DM. Through LASSO penalized logistic regression analysis combined with bootstrap resampling, a panel of 20 protein biomarkers that accurately predicted incident T2DM was identified with a significant incremental improvement over traditional risk factors. Finally, a Mendelian randomization analysis provided support for a causal role of 48 proteins in the development of T2DM, which could be of particular interest as novel therapeutic targets.

genetics↗