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Culture Chambers for Live-Cell Imaging
Specimen chambers are an integral and critical branch in the history of live-cell imaging, and a wide spectrum of designs have been published over the years describing systems that offer excellent optical properties while allowing specimens to be maintained for varying amounts of time. Ranging in complexity from the simple preparation of a sealed coverslip on a microscope slide to sophisticated perfusion chambers that enable tight control of virtually all environmental variables, culture chambers are designed to to allow living specimens to be observed with minimal invasion at high resolution.
Regardless of their design, live-cell imaging chambers must fulfill a variety of requirements in order to be successfully employed in experiments. The chamber should be easily sterilized and totally isolated from the laboratory environment with a cover or seal during observation to minimize exposure to sources of contamination. On the other hand, the culture chamber should also offer uncomplicated access to the cells if the investigation involves microinjection, addition of reagents (such as drugs or metabolites), physical manipulation of the cells, or changes to the culture medium. If the live-cell experiment is followed by cloning or fixing and staining the culture with synthetic fluorophores or immunofluorescence after the imaging session, the chamber must be configured to allow removal of the coverslip.
The culture chamber dimensions, including overall size, surface area for cell culture, and the volume of medium (or buffer) bathing the cells, are important considerations. In addition to being readily accommodated by the microscope stage, the culture chamber must be large enough in its lateral dimensions to contain sufficient cells for a thorough sampling of the population. The depth of the surrounding medium should be minimized to ensure the highest possibility optical quality for transmitted light and accessibility to the cells, yet it must also be deep enough to provide a healthy environment. Plastic coverslips should be avoided due to their inherent autofluorescence and strain birefringence, which interfere with some imaging modes. Only the highest-quality glass coverslips should be employed, and these should be cleaned with a strong acid or base and thoroughly washed. Commercial culture chambers vary in reliability, versatility, and cost. These factors should be considered when designing experiments in order to determine whether a single complex (and often expensive) chamber is required, or if a larger number of simple culture dishes will suffice. In the final analysis, it should be recognized that there is no perfect culture chamber for all purposes and many compromises are often necessary to achieve success in live-cell imaging.