Biology subjects

Ashman, K.

Publications and source records attributed to Ashman, K..

3 recordsLinked to original sources

bioRxiv2023

Heat n Beat: A universal high-throughput end-to-end proteomics sample processing platform in under an hour

Proteomic analysis by mass spectrometry (MS) of small ([≤]2 mg) solid tissue samples from diverse formats requires high throughput and comprehensive proteome coverage. We developed a near universal, rapid and robust protocol for sample preparation, suitable for high-throughput projects that encompass most cell or tissue types. This end-to-end workflow extends from original sample to loading the mass spectrometer and is centred on a one tube homogenisation and digestion method called Heat n Beat (HnB). It is applicable to most tissues, regardless of how they were fixed or embedded. Sample preparation was divided to separate challenges. The initial sample washing, and final peptide clean-up steps were adapted to three tissue sources: fresh frozen (FF), optimal cutting temperature (OCT) compound embedded (FF-OCT), and formalin-fixed paraffin-embedded (FFPE). Thirdly, for core processing, tissue disruption and lysis were decreased to a 7 min heat and homogenisation treatment, and reduction, alkylation and proteolysis were optimised into a single step. The refinements produced near doubled peptide yield, delivered consistently high digestion efficiency of 85-90%, and required only 38 minutes for core processing in a single tube, with total processing time being 53-63 minutes. The robustness of HnB was demonstrated on six organ types, a cell line and a cancer biopsy. Its suitability for high throughput applications was demonstrated on a set of 1,171 FF-OCT human cancer biopsies, which were processed for end-to-end completion in 92 hours, producing highly consistent peptide yield and quality for over 3,513 MS runs. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=152 SRC="FIGDIR/small/559846v2_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@4b8399org.highwire.dtl.DTLVardef@1acc573org.highwire.dtl.DTLVardef@1d7155eorg.highwire.dtl.DTLVardef@1bbef10_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2023

The impacts of contemporary logging after 250 years of deforestation and degradation on forest-dependent threatened species

Despite the importance of safeguarding forests and woodlands for achieving global climate and biodiversity agendas, logging continues across most forested countries. Forestry advocates often claim logging has minimal impacts, but rarely consider the cumulative threat deforestation and degradation has had, and continue to have, on species. Using New South Wales (Australia) as a case study, we quantify the extent of deforestation and degradation from 1750 - current. Using these estimates of overall loss as a baseline, we then quantify the relative extent of contemporary (2000 - 2022) logging and the condition of the remaining native forest and woodland (quantified by measuring the similarity of a current ecosystem to a historical reference state with high ecological integrity). Using these data, we measure the impacts on distinct vegetation types and on 484 terrestrial forest-dependent now-threatened species. We show that more than half (29 million ha) of pre-1750 (pre-European colonization of Australia) native forest and woodland vegetation in NSW has been lost. Of the remaining 25 million ha, 9 million ha is degraded. We found contemporary degradation from logging affected 244 forest-dependent now-threatened species that had already been affected by this historical deforestation and degradation, but the impacts varied across species and vegetation types. We found that 70 now-threatened species that were impacted by historical deforestation and degradation and continue to be impacted by logging, now have [≤]50% of their pre-1750 extent remaining that is intact (with three species now having <20%). By quantifying the historical impacts of deforestation and degradation, our research sets the impact of contemporary degradation from logging in perspective and highlights shortfalls in current environmental assessments that fail to consider appropriate baselines when reporting on overall impact. Future land management decisions need to consider not only the extent of remaining habitat based on pre-1750 extents, but also its condition. Article impact statementThe impact of logging needs to be placed in perspective by considering past losses and degradation due to human land use decisions.

ecology↗

bioRxiv2022

A Camera-Assisted Pathology Microscope to Capture the Lost Data in Clinical Glass Slide Diagnosis

Digital pathology, or the practice of acquiring, managing, and interpreting high-resolution digital images from glass pathology slides, holds much promise in precision medicine, potentially transforming diagnosis and prognosis based on computational image biomarkers derived from digital tissue images. However, for all its promise, digital imaging in pathology has not yet become an integral part of the clinical workflow as it has in radiology due to high cost, workflow disruptions, burdensome data sizes and IT requirements, and additional dedicated personnel requirements. Consequently, pathology retains the 150-year-old analog workflow, and the vast majority of slides used in clinical diagnosis are never digitized. Furthermore, there is a missed opportunity to capture the image information and associated data on search processes that led to the clinical diagnosis, which could serve as the foundation for computational clinical decision support. This paper describes an approach for slide digitization during clinical review using a camera attached to a standard brightfield pathology microscope. While a pathologist reviews a glass slide using the eyepiece oculars, the continuously running camera digitizes a complete record of the slide review, resulting in multi-resolution slide images and spatiotemporal saliency maps of the slide review. Unlike other approaches, the pathologist does not stop to review the video stream or monitor the acquisition of video frames but performs the diagnostic review at the microscope using the standard clinical protocol. This hybrid analog-digital approach combines the benefits of digital slide analysis, including annotation, computation, and the ability to confirm the completeness and quality of the glass slide review with the ease of using the microscope for primary diagnosis. Furthermore, a record of the pathologists attention during the review, including their search path, magnification level, and dwell times at each location on the slide, is obtained. In the future, this approach could enable the development and application of new and emerging computational decision-support algorithms in real-time to provide feedback to the pathologist, reduce diagnostic errors, and improve disease diagnosis and prognosis.

bioengineering↗