Cell Sorting Analysis by Flow Cytometry

Cell sorting is the separation and isolation of various cell populations. Flow cytometry sorters make physical separation of sub-populations of cells from a heterogeneous population, with a high degree of purity and high speed possible. This technique can be used in many situations, such as sorting sperm for subsequent insemination to allow sex selection of offspring, single-cell cloning of hybridoma cells for the production of monoclonal antibodies, purifying different lineages, including stem cells from bone marrow samples, and so on.

The advantages of cell sorting based on flow cytometry is that it is able to use multiparametric analysis to identify highly specific populations. Moreover, it is not just phenotypic characteristics identified by a specific antibody-antigen interaction that can be measured; it is also possible to measure the DNA content of cells, the RNA content, or even assess functional characteristics such as ion flux or pH oraltered cell states such as apoptosis and cell death.

Some methods can be used in flow cytometry cell sorting:

Mechanical sorting

Some flow cytometers cell sorter use a mechanical device called the catcher tube to sort the cells of interest. The catcher tube is located in the upper portion of the flow cell and moves into the stream to collect the cells. When cells pass through the laser beam the system determines if each cell belongs to the selected population defined by boundaries in the cytogram. If the cell is identified as a cell of interest, it is captured by the catcher tube and collected into a tube or into a concentration module, otherwise it is dispatched to the waste tank. The operator can choose the purity level of the sort among three levels (three sort modes).

Electrostatic sorting

The particles are ejected through a nozzle and forced to break up into a stream of regular droplets by applying a vibration to the nozzle. Droplets containing particles of interest can be deflected from the main stream by applying a charge to those droplets. When a charge droplet passes through a high voltage electrostatic field, between the deflection plates, it is deflected and collected into the corresponding collection tube. The deflection of the droplet is towards the oppositely charged plate, so that this droplet is separated from uncharged and oppositely charged droplets.

Other methods

Another way to sort cells is to use magnetic beads; it ispossible to positively select cells of interest by adding magnetically labeled antibodies to specifically select the population of interest or by negatively selecting a population by adding antibodies, coupled to magnetic beads, specific for cells other than those of interest. Cells are then passed through a column between a strong magnetic field to either elute or retard the population of interest.