Light sheet microscopy for live cell imaging of photosensitive systems

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Researchers at the Marine Biological Association (MBA) have a long standing reputation for excellence in the use of microscopy to study marine organisms. The ability to observe the unusual and novel physiology of these organisms has led to many significant discoveries. The new microscopy facility requested in this proposal will allow MBA researchers to examine cellular processes at unprecedented levels of resolution, with minimal stress on the cells or organisms under study, providing a world-class facility for future research into cell biology of marine organisms.Research at the MBA relies on advanced fluorescence microscopy techniques to study marine model organisms, using fluorescent dyes and proteins to view where specific proteins are found in a cell or to directly image cellular processes such as changes in intracellular pH or calcium, which are central to many signalling pathways. Other aspects of MBA research use model organisms to study how cellular processes have evolved or may operate in more complex multicellular organisms. For example, we use algae to study the structure and function of flagella, as many aspects of flagella are highly conserved with the cilia found in the human body. Marine phytoplankton are tiny algae that represent the base of the food chain in our oceans, supporting fishing and seafood industries, but despite their importance to life on our planet we still understand very little about them. Many phytoplankton surround themselves with intricate extracellular coverings made from silica or calcium carbonate. We aim to understand how and why these algae produce such intricate and ornate cell coverings to aid the design of similar structures for use in biotechnology and biomedical applications. Other research uses marine organisms from lineages that diverged at the very base of the animal tree of life, such as sea anemones, to examine how complex regulatory processes involved in embryo development evolved in multicellular animals. The marine fungi represent another group of organisms that are poorly understood. Recent research has highlighted that fungi are widespread in marine environments, suggesting that they are likely to play similar roles in nutrient recycling and as pathogens as the terrestrial fungi. We are developing model species of marine fungi for laboratory studies, so that we can understand much more about their physiology and their potential roles in the environment.Fluorescence microscopy of very small cells or structures requires that the subject is illuminated with a bright excitation light, which for many samples can be damaging. This is particularly relevant for photosynthetic organisms (such as marine phytoplankton) or for long term studies (such as time lapse imaging of biomineralisation or embryo development). MBA researchers are therefore currently limited in their ability to study important aspects of their model organisms. This proposal aims to address these limitations through the acquisition of a light sheet microscope. Light sheet microscopes are very effective at reducing damage from excitation light because they illuminate the sample with a very thin layer of light from the side, rather than passing light directly through the sample from above or below. Illumination from the side also has the effect of increasing the resolution of the microscope as there is no interference from out of focus areas of the sample. Light sheet microscopy has been shown to be particularly effective for long term time lapse imaging of photosensitive organisms for extended periods and therefore it is an ideal solution to the problems currently faced by MBA researchers.The new facility will be used to support the ongoing research of the current research teams, with interests from marine fungi and algae through to marine animals, and further afield. The resource will therefore underpin many diverse avenues of research and therefore represents excellent value for money.