Cell sorting fits into a special niche in the flow cytometry world. This technology enables users to isolate specific cell populations using cellular targets that can be stained with fluorescent antibodies. Cell sorting is a unique tool because it gives users the power to find and collect cells that are proverbial ‘needles in a haystack.’ Many flow cytometry users are reluctant to take on cell sorting because they are wary of using an instrument they view as complicated and finicky. But most cell sorters today are very user friendly and don’t invoke the use of oscilloscopes and black magic like their first generation counterparts.

New alternative technologies, like magnetic bead separation, have given scientists an alternative to cell sorting for some routine cell separation protocols. Magnetic beads can’t handle every cell separation, and there will always be a place for sorting.

• Assure that the cells you want to sort are stained with a panel that allows for maximum separation

• Have a good understanding of your cell sorter: Make sure the machine is properly set up, the fluidics are stable, and regular QC (quality control) procedures are in place and within acceptable ranges 

Many flow cytometry core facilities have a dedicated expert that runs the cell sorter. This person is a source of vital information about the instrument and its capabilities, and you should consult with her or him as you design your cell sorting experiment. Experimental considerations should include your panel design and estimating how many cells you need to start with in order to get the desired number of sorted cells. Cell sorting can result in the loss of a lot of cells along the way, especially if you are dealing with a population that is not easily separated.  So it’s worth maximizing the number of cells you start with in order to aim for a reasonable sort.

Cell sorting is uniquely suited for certain applications like isolating single cells with a particular phenotype that can be clonally expanded into a pure population. Antibody-expressing cells can also be sorted, which can save significant time for individuals screening cells for hybridoma fusions. So give sorting a chance and confront your irrational cell separation fears!