Current Diploma Projects
Below you can find out the work that is being undertaken by our current RMS Diploma students:
Developing protocols and training materials for the live imaging of Drosophila via lightsheet and multiphoton microscopy
Jennifer Adcott, University of Liverpool
Lightsheet and multiphoton microscopy have been widely used to reduce photo-damage. Here these systems will be used and compared for the imaging of live Drosophila using established cancer model fly lines. A protocol will be established for sample preparation, image acquisition, through to image processing and analysis. Suitability for each technique will be compared for addressing specific questions where live imaging of small model organisms or ex vivo tissues is required. The aim will be to publish these results as a methodology paper with JoVE video protocol.
Elucidating the artifacts and abberations of SIM and STORM super resolution imaging technologies and subsequent protocol optimisation for cells and tissue sections
Julian Mark Bowen, University of Cambridge
Once our super resolution system is installed I will attempt to identify the reasons for artifacts and aberrations within the images and develop and optimise methods to eliminate them. Sample preparation is known to be critical to obtain good images using widefield and confocal imaging but is especially important for super resolution imaging.
The Characterisation of Speciality Fibres Utilising 3-D SEM Constructed Images
Michael Brookes, University of Leeds
Distinguishing expensive speciality animal fibres such as cashmere and alpaca from cheaper fibres such as wool is an important activity. Current techniques, such as ISO 17751-2, measure fibre diameter utilising an SEM in 2 dimensions at high vacuum and susceptible to viewing errors. The study will use 3-D imaging techniques to measure and characterise speciality fibres and compare the results with that of current techniques.
Comparative 3D imaging of gene expression in Arabidopsis
Maria Conejero Moreno, Royal Botanic Gardens Kew
Immunolabeling and in-situ hybridization have been used to study the location of genes involved in plant development in 2 dimensions. For my diploma I propose to extend this methodology, developing a method for the visualization in 3 dimensions of gene expression in plant tissues using phase contrast X-ray radiation microtomography. This work should increase the understanding how genes are involved in plant function and morphology during different stages of growth. Using Arabidopsis as a model system the project will shed light on the spatio-temporal expression of genes during floral development.
Testing, optimising and implementing cryo-sample preparation and imaging techniques to optimise sample quality and acquisition efficiency in single particle analysis
Adam Costin, University of Oxford
With the rise of cryo-EM as a powerful tool for solving protein structures there has been an influx of new researchers learning the technique. Sample optimisation for cryo-EM remains slow and highly inconsistent. This study will aim to create a workflow that will systematically optimise sample preparation much faster for new users. This will be done through a comprehensive assessment of current techniques and then testing these within the context of an EM Facility. A way of simply incorporating tomography into the process will also be developed, as this is a powerful way of assessing ice thickness and protein structure.
The Study of Marek’s Disease Virus In Feather Follicle Epithelium
Jennifer Simpson, The Pirbright Institute
Marek's disease virus (MDV) is an alphaherpesvirus which causes a lymphoproliferative disease in chickens. Inflection occurs by inhalation of infected dander and following a complex life cycle involving infection of B and T cells, the virus is shed from the feather follicle. Vaccination is available and prevents clinical disease however it does not prevent the shedding of virus. The feather follicle is important in the transmission of MDV as the feather follicle epithelium (FFE) is the only region where fully enveloped and infectious virus particles are assembled. This study aims to identify cell tropism and describe viral morphogenesis in the FFE using immunofluorescence and transmission electron microscopy.
Comparative Analysis of Cellular Localisation of Viral Glycoproteins and Capsid Protein in Transfected Cells
Helen Todd, The Moredun Research Institute
A9.5 is a gene unique to the wildebeest-associated MCF virus, Alcelaphine herpesvirus-1. A Haemagglutinin (HA) epitope-tagged construct has been generated that can be transfected into HEK293T cells and has been found to express the A9.5 protein by Western blot. A9.5 encodes a predicted glycoprotein with a potential N-terminal signal sequence but the protein sequence has no similarity to known proteins. In order to define the cellular localization of A9.5 we will compare it with the known cell-surface viral antigen glycoprotein B (gB) and the major virus capsid protein (ORF25), by transfection of HEK293T cells and analysis by fluorescence microscopy.
Characterisation of the Migratory Properties of Additives used in Acrylic Coating on Polymer Film
Craig Holliday, Innovia Films
Polypropylene film is often coated with acrylic nano-particle formulations which contain migratory additives designed to modify the surface properties of the material. Whilst it is possible to quantify the levels using infra-red spectroscopy, this only provides a bulk assay.
It is hoped that a method to image the dried acrylic coating and map additive distributions on the dried surface can be developed to allow a study of the interaction of bulk levels and drying temperature on the migratory behaviour. This can then be correlated to physical and optical characteristics to optimise manufacturing conditions.
First of all, the RMS Diploma was fun, I have nothing but good things to say about it and I learnt a lot of microscopy and biology. The project I undertook for the Diploma was a cell death project (how to measure necroptosis.) The RMS Diploma has helped me a lot, opening several doors for me. I was able to get my name on 5 publications because of it and gave a talk at the flowcytometryUK meeting this year. I also helped out with a workshop on cell death at the same meeting.
Since the RMS Diploma I have taken over the light microscopy facility at ICR and merged it with the flow cytometry facility. The facility is now called “Flow Cytometry and Light Microscopy Core Facility”. This increased responsibility was a direct consequence of doing the RMS Diploma in my opinion. As I am now running the microscopy facility I was able to attend the AQLM course in Woods Hole this year. It is a great 10 day course on microscopy which enhanced my knowledge of microscopy a lot. I am very grateful to the institute for supporting me with this course since it is a large cost to attend it, but I think finishing the Diploma showed the ICR that I am serious about learning microscopy and working in the field which led to me acquiring the funding. We are also investing a lot in microscopy this year at ICR which will grow the facility and give me a lot more responsibilities.
I would say that a lot of these new responsibilities and opportunities in my working life have been made possible because I completed the RMS Diploma.