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Cell Sorting
Cells can be sorted in a sterile environment enabling the recovered cells to be cultured. This is especially applicable to transfected cells, where only a small proportion of the cells may be expressing the antigen of interest. We are able to sort according to antigen expression, GFP expression, DNA content or cell function (eg calcium flux or apoptosis). More information about the theory of sorting can be found here.
We can also sort under non-sterile conditions to retrieve sub-populations for RNA and protein extraction or for morphological analysis. Cells can also be sorted singly into 96 well plates (or the receptacle of your choice - within reason) for cloning or PCR. Sterile and non-sterile sorting is performed on the FACS Vantage or the MoFlo.
The sorting can be optimised according to whether purity, recovery or count accuracy is most important.
Sort rates will vary between the sorter used and the cell type being sorted. As a general rule of thumb, on the FACS Vantage we can run cells at about 4,000/second or about 14e6/hour. On the MoFlo, we usually run at about 15,000/second or 54e6/hour. In general, small round cells will be able to be run faster than epithelial cells.
The default nozzle size on both cell sorters is 70µm; for larger cells, we can use a 100µm nozzle but this will also result in a slower sort speed. Further details about the practicalities may be found on the Local Users Page (available to CR-UK researchers only) or by emailing one of the staff of the Lab.
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IMPORTANT
Only Hazard Group I (Advisory Committee for Dangerous Pathogens) / Class
I genetically modified organisms, as defined by the Genetic Modification
(Contained Use) Regulations 2000, will be accepted for cell sorting in the
FACS Laboratory.
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