Abstract
We share two simple modifications to enhance the fixation and imaging of relatively small, motile, and rounded model cells. These include cell centrifugation and the addition of trace amounts of glutaraldehyde to existing fixation methods. Though they need to be carefully considered in each context, they have been useful to our studies of the spatial relationships of the microtubule cytoskeletal system.
Highlights
The striking mobility of Dictyostelium makes for an interesting and impactful model system to study dynamic cellular events [1]
We have tinkered with fixation and staining conditions in order to image cytoskeletal arrays and GFP-tagged proteins in D. discoideum [2,3,4,5,6]
Cells under agarose can undergo division and remain viable for hours in sealed chambers. This overlay remains our preferred method for live cell imaging, enabling us to follow individual microtubule (MT) motions in a minimal number of focal planes [8,9]
Summary
The striking mobility of Dictyostelium makes for an interesting and impactful model system to study dynamic cellular events [1]. One of the tradeoffs in a dynamic lifestyle is that these cells can be less tightly adherent to their substrate, and imaging and downstream structural analyses can present challenges that must be carefully considered. We have been challenged with difficulties in getting cells to sufficiently flatten to facilitate imaging and to remain attached to coverslips for downstream structural processing. In this brief report, we would like to share two insights that make a difference in our work, in the hopes that they benefit others as well
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