BioLaser: Establishing a High-Resolution Laser Ablation Tomography Platform for UK Bioimaging Research

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Background summaryTwo- and three-dimensional (tomographic) images of biological materials provide insight into the relationships between subcellular structure and function. However, particularly for hard or opaque tissues, instruments that can do this are currently large and extremely expensive. Furthermore, sample preparation and/or imaging is slow, preventing cost effective analysis of the many samples commonly required for biological studies. Thus, there is urgent need for new methods to image and quantitatively describe biological samples at high resolution and speed, allowing analysis of traits currently considered intractable to high-throughput investigation. While the component laser and imaging technologies exist, getting them to work together, and crucially, to achieve high-resolution sub-cellular imaging on biological samples, requires modifications and early concept exploratory and validation investigations. We propose novel modifications to existing, first-generation laser ablation (high-powered, ultrafast laser pulses) technology, enabling new biological insight and understanding into biological problems that were previously intractable using conventional techniques.Project summary'BioLaser: Biological Investigations by Laser Ablation Tomography' is a pump-priming project that aligns world-class expertise in laser technology (Institute for Manufacturing, IFM, at the University of Cambridge), plant research, physiology (NIAB, Cambridge) and image analysis (both centres). The approach centers around the use of precision lasering (based on a technique called 'Laser Ablation Tomography', LAT). This will be combined with in-process monitoring to adjust the ablation process according to the composition and topography of the sample ('metrology'), to generate precision ablated biological samples of unprecedented quality for microscopy and 'chemical imaging' platforms that render high-resolution 2D & 3D images, allowing quantitative analysis of cellular and subcellular structures. Using an array of plant tissues, we will evaluate & validate the parameters required to design a precision LAT bioimaging system. This project is structured around for key approaches, all of which must be addressed to successfully develop LAT-based methodologies for high-resolution bioimaging:1. Expertise in laser-based manufacturing technologies: IMF provides this project with world-class expertise in precision laser research, engineering and manufacturing. Such on-site expertise is the foundation for project success.2. Precise ablation control: To achieve high-resolution imaging, a uniform plane is required. This needs precise knowledge of the position and power of the laser alongside measurement of the ablated surface after each laser pulse, provided via in-process metrology to ensure high-resolution imaging is truly achievable.3. Advanced tools for imaging and analysis: The precision machined samples produced will be investigated using a suite of state-of-the-art 2D & 3D imaging and analysis approaches, exploiting imaging expertise at the UofC.4. Image validation: the tools and approaches to be developed are powerful and precise. However, used without parameterisation and validation, the resulting images could be of limited scientific value. The image data generated will be validated using conventional approaches, providing a feedback loop for parameter optimisation.This project will lay the foundation for future manufacture of a benchtop prototype, delivered via IFM. Achieving the project aims would result in a step-change over first generation LAT systems that currently work at low magnification. Ultimately, we aim to establish a LAT bioimaging hub based in Cambridge, and to facilitate the production of similar platforms for the UK and beyond. This capacity-building approach will promote the critical-mass in technology uptake necessary for the potential impact of precision LAT bioimaging to be fully realised.