After the fixation/permeabilization step the cells are gone. Where did they go?
The Fixation/Permeabilization is not a lysis step but rather a fixation step that also permeabilizes. It was optimized for staining with freshly isolated PBMCs or mouse splenocytes. The cells become smaller and are more translucent giving the appearance that the pellet is greatly diminished. But if the pellet is resuspended, the same number of cells will be present as confirmed by observing the cells under the microscope. Because of the change in size, the voltages have to be adjusted to “find” the cells. We have tested many other buffers and have not found any commercially available buffer to give the same robust and reproducible results as the Foxp3 buffer we supply.
Related Questions
- How long can cells that have been stained with eFluorTM 605NC and eFluorTM 625NC conjugated antibodies followed by fixation be stored prior to analysis?
- How long can cells that have been stained with eFluorTM Nanocrystal conjugated antibodies followed by fixation be stored prior to analysis?
- How long is the fixation step?
