Sorting and aggregates tip sheet

Aggregates or clumps can severely affect the integrity of your sort no matter which instrument you work with.

Clumps cause clogs, which block or disturb the laminar flow process through the nozzle. Such disturbance usually results in redirection of the central core stream or spraying of the side streams, which affects the purity of the sort.

While we can manually manipulate the instrument to counteract some of the hazards of aggregates, three bench-based steps can help:

1. Use PBS or HBSS without calcium and magnesium as the base of all the solutions you need in preparing cells for sorting. In some cases 1 mM EDTA is also recommended.
2. Keep the protein concentration in your sort buffer to 0.5% (FCS or BSA). Collection vessels can have unlimited protein to keep cells happy once they’ve been sorted.
3. Filter samples immediately prior to sorting by passing them through 50–70 μm nylon mesh.