Flow Cytometry: Basic Definitions

A filter that transmits light wavelengths within a selected range

The point at which droplets break from the stream

Method by which particles are analyzed in a liquid stream that is subsequently partitioned into droplets that are deflected and collected based on scattered light and fluorescence selection criteria

A glass filter that transmits light of a particular wavelength range in the forward direction but deflects light outside of the range at a 90° angle

A graph displaying two measurement parameters (scatter, time, or fluorescence) plotted on an X-Y axis

Two particles that pass through the laser so closely they are recorded as a single pulse or event; doublets can be resolved using pulse area in combination with pulse width or height

The distance between the interrogation the point and point at which the first droplet breaks off (the break-off point)

The wavelength of maximum emission of a fluorescent molecule

An electronic pulse, generally caused by a particle passing through the interrogation point, that is converted to data representing a single particle

The wavelength of maximum absorption of a fluorescent molecule

A technology used to analyze single particles in a liquid stream by measuring their physical and chemical properties as they pass through an excitation source. The resulting emitted and scattered light are detected and measured

The rate at which the sample flows through the cytometer

The mathematic process of correcting for the detection of photons from the spillover of a fluorophore into multiple detectors

A compound that can re-emit light upon excitation

Light scattered in the forward direction, typically 180° from the laser beam axis ; this measurement correlates with a particle’s relative size

User-defined region drawn on plots to select a population of events and exclude them from other events in subsequent analysis

A graph displaying the distribution of the number of events in one parameter of measurement (scatter or fluorescence)

The point at which cells enter the illumination of the laser

A light source that produces a single wavelength of light at one or more discrete frequency

A glass filter that transmits light wavelengths longer than a designated wavelength

A signal pulse generated in a detector that reflects the passage of a cell or particle through the laser beam characterized by height, area, and width

Height: The maximum amount of current output by the photo multiplier tube (PMT)
Area: The integral of the pulse
Width: The time interval during which the pulse occurs
Signal intensity can be measured by either height or area
The width parameter measures the time that the cell spends in the laser

Quantifying a pulse by measuring its height, area, and width.

The number of sorted particles of interest compared with the total number of sorted particles

Light scattered at the interrogation point typically at a 90° angle to the laser beam axis; this measurement correlates with relative cell complexity or granularity

The process of converting photons emitted by fluorescent dyes and probes into a digital pulse, which can be integrated and converted into a numeric value that can then be graphed and analyzed

A single particle that passes through the laser and is recorded as a single pulse or event

Light that is emitted by one fluorochrome but measured in the detector designed to detect another

The difference between a fluorophore’s excitation maximum and emission maximum

A value set in the software that triggers the electronics to ignore background signals that are under it

The total number of particles of interest sorted compared to the total number that could have been sorted under ideal conditions