Dr Gemma Cairns - The Development of M4All: MultiModal Modular Microscopy for All
M4All is a fully open-source and 3D-printable optics system which can be used to build low-cost microscopes combining different imaging modalities. M4All was primarily developed to address imaging challenges when investigating macrophage-pathogen interactions. Streptococcus pneumoniae is the most common cause of community acquired pneumonia. Mitochondrial reactive oxygen species (mROS) have been shown as critical microbicidal factors employed by alveolar macrophages in the clearance of internalised bacteria. Understanding and quantifying interactions with mROS could provide a future target for pharmacologically enhancing host responses to S. pneumoniae as an alternative to conventional antimicrobials, to address the increasing issue of antibiotic resistance. This is especially important in some developing countries in Africa and Asia where prevalence of invasive pneumococcal diseases is much higher.
The short half-life of mROS (from 10-9 s to a few seconds) and the time frame of the overall response of alveolar macrophages to S. pneumoniae (16+ hours) poses challenges with detecting short lived interactions over long time frames. Advanced microscopy hardware for live cell microscopy can be expensive, leading to issues with accessibility. Furthermore, imaging over long periods of time introduces challenges with photobleaching and phototoxicity. Low-cost microscopes have the potential to widen accessibility to these techniques. With a focus on stability for timelapse imaging studies within incubators, M4All was developed as modular CAD files which are printed monolithically to build up the desired optomechanical system. M4All is also compatible with the OpenFlexure microscope stage. We have developed three microscopes using M4All in combination with the OpenFlexure microscope: a dual channel fluorescence microscope, a single channel fluorescence and computational phase contrast microscope, and a single channel brightfield incubator microscope. Here I present initial imaging results of macrophages and discuss how M4All could enable wider accessibility to advanced technologies by combining low-cost 3D-printable microscopes with computational microscopy techniques.
Dr Fernan Federici - How open source hardware projects have enabled our capabilities for education, research, and outreach.
The need to address inequalities in education, scientific research, and technological development has brought open science and open source technology to the forefront. The use of open protocols, public domain reagents, and open hardware is leading to more inclusive, efficient, and multidisciplinary research models. Collaborative networks such as ReClone, JOGL, iGEM, and GOSH, which openly share these resources online, are also enabling the emergence of more decentralized models for biotechnology. In case of Dr Federici and his group of scientists, engaging with these communities has expanded their research opportunities and facilitated the development of new teaching resources. For instance, they were able to use well-documented open hardware projects, such as OpenFlexure, OpenLabTools, and FlyPi, to create microscope prototypes for various applications, ranging from yeast monitoring during fermentation processes to time-lapse fluorescence experiments with optogenetic control. During the COVID-19 lockdown, they assembled portable minilabs for teaching hands-on molecular biology practicals at home. In this presentation, Dr Federici will share their experiences working with open source hardware projects in his lab and within the context of the local reGOSH-CYTED Latin American network. Furthermore, he will discuss the limitations they are facing in expanding these open frameworks in our region and share insights into how the plan to address these challenges.
Dr David Baddeley - Python-microscopy and pyoptic - open source software for microscope control and design
Python-microscopy is a suite of software tools for the control of open-source microscopes, for simulating microscopic imaging, and for image data analysis. Dr Baddeley will introduce python-microscopy, describe some of the innovations they have made for handling high data-rate streams and extended imaging durations, and discuss how to interface with custom hardware. He will also describe the use of pyoptic2 – a python-based optical-CAD package – in conjunction with python-microscopy and 3D printing for the simulation and rapid-prototyping of custom optical systems.
Dr Rifka Vlijm - Going Live! Technological developments for live cell (STED) super-resolution imaging
With the development of super-resolution microscopy, and great progress in labelling techniques, direct visualization of cellular structures in living cells now is possible at resolutions of 30-40nm. Commercial Stimulated Emission Depletion (STED) microscopes now ensure access to this technique to researchers not specialised in optics. One important limitation which hinders a broad application to cell biological questions is the low throughput due to the manual steps required for optimal results still. Capturing ‘rare’ events, such as for example a specific cell division stage, with only manual structure selection and image parameter selection becomes near impossible. To overcome these limitations, Dr Vlijm and her group have managed to fully automate our data acquisition, and thereby improved the throughput by two orders of magnitude. They have shown the value of these improvements by detecting cells in a specific cell division stage without the typical required chemical or genetic modifications necessary to increase the occurrence. In combination with full incubator conditions on the microscope the scientists have furthermore developed a protocol for live cell STED which still enables normal cell proliferation.
