Flow cytometry antibodies, kits, reagents

Chapter 1: Principles of the Flow Cytometer

Fluidics System

One of the fundamentals of flow cytometry is the ability to measure the properties of individual particles. When a sample enters a flow cytometer, the particles are randomly distributed in the 3-D space of the sample line, the diameter of which is quite a bit larger than the diameter of most cells. The sample must therefore be ordered into a stream of single particles that can be interrogated individually by the machine’s detection system. This process is managed by the fluidics system.

The fluidics system consists of a central core through which the sample fluid is injected, enclosed by an outer sheath fluid. These are pushed through the system at slightly different pressures. The sheath fluid is under high pressure and, therefore, moves faster. As the sheath fluid moves, it creates a drag effect that causes the sample in the central core to narrow (Figure 1). This creates a single file of particles and is called hydrodynamic focusing. Under optimal conditions (laminar flow) the fluid in the central chamber will not mix with the sheath fluid.

Hydrodynamic focusing produces a single stream of particles.

Figure 1: Hydrodynamic focusing produces a single stream of particles

Without hydrodynamic focusing, the nozzle of the instrument (typically 70 or 130 µm) would become blocked. The cells flow in single file through the illumination source, allowing them to be analyzed one cell at a time.