MACSQuant TytoCartridge-based cell sorter
The MACSQuant Tyto is built around the idea of keeping the sample separated from contamination sources – or, inversely, keeping the sample from contaminating anything else. Cell sorting on a MACSQuant Tyto occurs entirely in a closed cartridge that is divided in to three chambers: the input, positive collection, and negative collection.
In the comfort of a sterile tissue culture flow hood, the user loads the sample in to the sample chamber. Once the cartridge is sealed, nothing goes in or out of it until the sort is complete. The user can then bring the cartridge to the instrument and install it in the sample block.
To sort the cells, low air pressure (< 3 psi) is applied to the sample, which is then pushed through a fluidic chip. The cells are interrogated by the lasers, the signals are processed, and cells within the target population are diverted into the positive chamber by a high frequency valve. The unsorted flow through continues towards the negative chamber.
Fragile samples will fare better sorting at low 3psi pressure (for comparison, low pressure on the Aria is ~30psi)
The MACSQuant Tyto is equipped with three 100mW lasers and 8 fluorophore detectors, plus the SSC and backscatter detectors:
488nm: 525/50, 585/40, 693/38 , 750LP
405nm: 450/50, 510/50
635nm: 693/38, 750LP
For best results, it is ideal to assign to highly expressed markers with very distinct on/off pattern of expression. Signal emission and light collection is impaired by the cartridge. B4 and R2 (PECy7 and APCCy7) are not suitable for dim marker and should be used for bright, primary markers. Bright fluorophores on better channels such as V1, V2, B1 and B2 (BV421, BV510, BB515 and PE respectively) are preferred to resolve secondary and tertiary markers. Note that FSC and SSC will not be appropriate for defining populations of cells, they must be defined/gated by a fluorescent marker.
Sort purity is a function of cell concentration on the Tyto
Once the sort is completed, a purity check should be performed to assess the result. Running the collected cells in a new cartridge is not a viable solution. We recommend sampling the collected cells on a benchtop instrument that will provide a true cell count, such as the Attune NxT or the Penteon.
For best result, you should run:
- a fraction of the sample
- a fraction of the collected cells
The Attune NxT and the Penteon will provide a value for your cell concentration and you will be able to know how many cells have been collected. Purity of the sort will be assessed but the display of your cells on the graphs: a pure sort should give tight clusters of cells with few (if any) outliers.
Unassisted cell sorting
- Charged based on time used
- The instrument will run 4mL/hr – mind the cell concentration for best results
- Staff time will be charged for initial template settings
- Once a template is designed, the instrument can be used without involvement from the staff
- Cartridges are available from the CAT facility at a discounted price
- Cartridges should be single use only