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Why live cell imaging?To understand the functional integrity of a whole liveing organism, it is important to first have a clear understanding of the interactions between both individual cells and single molecules within these cells. Using the latest in biomolecular / celluar imaging technology, researchers are focusing on investigating such biological interactions at both the molecular and cellular levels. This involves direct imaging of the "presence" of single molecules and individual cells as well as the use of real-time imaging of dynamic, reversibile biological signaling "events" such as protein-protein interaction, protein modification and intracellular calcium changes. Moreover, this imaging technology can be combined with protein manipulation techniques to elucidate the functional cotribution of these dynamic interacions and signal trasduction events in the whole living organism. Environment for the cell on the microscope 1. Temperature Most mammalian cells grow at 35-37 o C. If the temperature is not optimally controlled, cell growth can be largely compromised. High temperature is even more detrimental since it results in immediate cell death. Therefore, maintaining the temperature within an optimal range is very important for cellular imaging. When imaging with a high numeric aperture lens it is necessary to control the temperature of the objective also, as the optical coupling of the medium (oil, glycerin or water) acts as a thermal coupling medium, drawing heat away from the specimen. 2. pH correct pH is crucial for cultured cells to maintain their normal physiological states and thus remain viable over the long-term. The pH of the medium should be maintained around pH 7.4 In a bicarbonate buffer system, which is the most widely used buffer system in cell culture, pH depends on the level of CO₂. Thus, the CO₂ level should be maintained during cell imaging. 3. Humidity All incubator systems should maintain internal humidity to prevent medium evaporation. Evaporation causes the accumulation of cell metabolites and medium constituents that will eventually impair cell growth. |
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