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Live Cell Imaging
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Why live cell imaging ?

In order to understand the functional integrity of a whole living 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 focused on investigating such biological interactions at both the molecular and cellular level. This involves not only direct imaging the “presence” of single molecules and individual cells, but also the use of real-time imaging of dynamic, reversible biological signaling “events” such as protein-protein interaction, protein modification and intracellular calcium changes. Moreover, this imaging technology will also be combined with protein manipulation techniques to elucidate the functional contribution of these dynamic interactions and signaling transduction events in the whole living organism.
Live Cell Instrument (LCI) is an innovative company that designs, develops and manufactures live-cell microscopy environmental control systems for qualitative and or quantitative microscopy. Our products are dedicated to the science of live-cell microscopy and include imaging systems, micro-observation environmental control instruments, and specialized fittings and adapters as well as numerous supplemental items.

Environment for the cell on the microscope

1. Temperature
Most of mammalian cells grow at 35-37℃. If the temperature is not optimally controlled, cell growth would be slowed or metabolic imbalance would occur. High temperature is even detrimental to cells since it causes immediate cell death. Therefore, controlling the temperature within optimal range is most important factor for cellular imaging.

Time-lapse imaging of cultured hippocampal neurons. EGFP-tagged target protein is moving into dendritic spines by neuronal stimulation.
Chamlide EC-S-10/ HP-R-20 heating plate /IL-H-10 fluidic inline heater Dr. Sunghoe Chang, Department of Life Science, GIST
2. pH
The correct pH level is vital for both the long-term viability of cultured cells and maintaining their correct physiological state. The medium should be maintained around pH 7.4. In a bicarbonate buffer system, which is most widely used buffer system in cell culture, pH depends on the level of CO₂. Thus CO₂ level should be maintained during cell observation.

3. Humidity
Closed chamber system which doesn’t use perfusion, requires maintaining humidity to prevent evaporation. Evaporation caused the concentration of cell metabolites as well as medium constitutes which will impair cell growth.

Types of observation chambers

1. Perfusion chamber
Perfusion chamber circulates the media through installed inlet and outlet holes installed. Perfusion enables the experimenter to provide a fresh medium and to change the composition of medium as needed. With a fluidic inline heater (IHS-101), one can prewarm the perfusion medium, thus maintain the temperature of medium before it exposes to cells.

2. Closed chamber (Non-perfusion type)
Closed chamber does not use perfusion thus it requires the controlling of CO₂ level as well as humidity. Closed chamber is used when the experiment requires a long-term observation in a completely air tight environment after a certain treatment.
Time-lapse imaging of membrane tubule formation in CHO cells.
Membrane tubular structures are forming by overexpression of EGFP-tagged target protein into the cells.
Chamlide CF-S-10 /HP-R-30 heating plate /IL-H-10 fluidic inline heater
Dr. Sunghoe Chang, Department of Life Science, GIST

Why observation chamber with cover slip?

According to Abbe’s theory, the resolution of light microscope fully depends on the N.A. of the objective lens. Therefore, for high resolution microscopy, it is absolutely required to use high N.A. objective lens which unavoidably has very short working distance. The conventional plastic culture wares are not thin enough to use with a high N.A. lens, thus one should use a cover slip which has thickness over the range 0.15-0.20 mm. Besides, a cover slip doesn’t emit autofluoroscence and has an appropriate reflective index (~1.515) which is optimal to use with oil or water immersion objective lens.

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Calcium influx/efflux
Cell locomotion
Mitochondrial potential
Protein translocation
GFP/RFP expression
Cell mitosis
Zinc translocation