Monitoring cell retention during the cell freezing - thawing cycle. U937 cells, labeled with the Hoechst dye (blue fluorescence) and Quantum Dots (red fluorescence), were loaded into the i3C and then underwent a freezing and thawing cycle, which included the following documentation steps: first, before closing the cryostage lid; (A) transmitted light image is superimposed with the fluorescence image of the same field (B). Next, images were taken after closing the cryostage lid (C) and following the pre-cooling step (D), after which freezing was performed. Immediately after thawing, while the lid was still closed, transmitted light (E) and fluorescence (F) images were acquired. To assess the vitality of the same thawed cells, we examined their ability to hydrolyze FDA by exchanging their cryo-media with FDA staining media: 7.5 μm of FDA staining solution was introduced to the conduit basin of the i3C (when the i3C was in the cryostage chamber), and then (after a few minutes) the fluorescence of the FDA-stained cells was imaged (G). Note that the majority of cells retain their spatial location. The red arrow indicates a single cell that changed location after thawing. Bars: 25 μm.
Deutsch et al. BMC Cell Biology 2010 11:54 doi:10.1186/1471-2121-11-54