Biology subjects

Greenan, G.

Publications and source records attributed to Greenan, G..

4 recordsLinked to original sources

bioRxiv2026

Characterization Standard for In-situ Cryo-electron Tomography

Standardized biological specimens are essential for optimizing cryoEM workflows and benchmarking instrument performance. While apoferritin fulfills this role for single-particle analysis, no equivalent exists for cryo-electron tomography. Ribosomes are frequently used but require large datasets due to C1 symmetry and structural heterogeneity, limiting rapid optimization and standardized comparison of workflows. Here, we present PP7 virus-like particles (VLPs) overexpressed in E. coli as a scalable in situ benchmark. VLPs have high orders of symmetry enabling rapid, high-resolution validation of tomographic pipelines from minimal datasets, while their distinct structural features across low to high resolutions provide a practical resolution metric.

cell biology↗

bioRxiv2026

High Accuracy Fluorescence Guided Focused Ion Beam Milling

Cryo-electron tomography (cryo-ET) is a powerful approach for visualizing macromolecular structures directly within cells, but its broader application is limited by the difficulty of reliably targeting specific structures during sample preparation. In particular, capturing small or rare objects within cryo-focused ion beam (cryo-FIB) milled lamellae remains a major bottleneck. Here, we present two fluorescence-guided cryo-FIB milling workflows that overcome key sources of targeting error and enable routine capture of structures across a broad range of spatial scales. For larger targets (>500 nm), we present an improved registration-based strategy combining FIB-milled fiducials with depth correction for refractive index mismatch. For smaller targets (150-500 nm), we implement real-time fluorescence guided milling on a commercially available cryo-FIB-SEM platform enabling consistent recovery of small, low-copy organelles such as centrosomes and primary cilia. Both workflows are implemented in a new open-source package fibsemOS and together expand the range of cellular structures accessible to cryo-ET and establish a broadly deployable framework for targeted in situ structural biology.

cell biology↗

bioRxiv2025

Rehoming laboratory rats: Exploring perceptions of rehomers, animal technicians and biomedical researchers

Laboratory animals can be rehomed at the end of a study. However, there is relatively little research on stakeholders perceptions of rehoming. We explored the views of three key groups: the people who volunteer to rehome these animals, the animal technicians who care for these animals prior to rehoming, and the researchers who are normally responsible for most of the interactions between people and these animals. To do this, we carried out a thematic analysis of semi-structured interviews. Our aim was to obtain insights that could inform reflection and guide future research priorities around rehoming programmes. Our study demonstrated support for laboratory rodent rehoming in all stakeholder groups. Rehomers and technicians comments focused chiefly on the potential benefits of rehoming to the animal and benefits to institutional openness. Researchers made similar comments, but in this group, these comments were tempered by concerns around the welfare of the animals after rehoming, the fate of animals that cannot be rehomed, rehomers health and well-being, and the potential risks associated with the transparency of individuals and the institutions use of animals in research. We discuss these themes in the context of the ethics of laboratory animal use, manifesting a Culture of Care, and challenges around enhancing institutional openness on animal research.

scientific communication and education↗

bioRxiv2022

AreTomo: An integrated software package for automated marker-free, motion-corrected cryo-electron tomographic alignment and reconstruction

AreTomo, an abbreviation for Alignment and Reconstruction for Electron Tomography, is a GPU accelerated software package that fully automates motion-corrected marker-free tomographic alignment and reconstruction in a single package. By correcting in-plane rotation, translation, and importantly, the local motion resulting from beam-induced motion from tilt to tilt, AreTomo can produce tomograms with sufficient accuracy to be directly used for subtomogram averaging. Another major application is the on-the-fly reconstruction of tomograms in parallel with tilt series collection to provide users with real-time feedback of sample quality allowing users to make any necessary adjustments of collection parameters. Here, the multiple alignment algorithms implemented in AreTomo are described and the local motions measured on a typical tilt series are analyzed. The residual local motion after correction for global motion was found in the range of {+/-}80 [A], indicating that the accurate correction of local motion is critical for high-resolution cryo-electron tomography (cryoET).

cell biology↗