In the original paper by Skehan et al., and the nature protocol by Vichai et al., they fixated their cells with TCA, without aspirating medium beforehand. In other papers (e.g. Papazisis et al. 1997), an optimized protocol involves aspiration before fixation to increase the signal-to-noise ratio. However, this aspiration is performed with a microplate multiwash device to ensure minimal loss of cells. If such a device is not available, is it still better to aspire medium before fixating cells? In my experience, the cell loss can be quite significant when one removes medium by flipping the plates instead of using a multiwasher... Any suggestions or insights on this?
If you carefully read the paper of Sekhan et al or even Vichal et al, you will note the conditions for their work and see why did they not aspirate the cell suspension. On the other hand Papazisis et al did use the aspirating medium to increase the signal to noise ratio which was required for his work. Now it is up to you to decide what you want.
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