Flow cytometers are laboratory instruments that are used to analyse the physical and chemical characteristics of particles in a fluid as they pass through at least one laser.
Cell components are fluorescently labelled and then excited by the laser to emit light at varying wavelengths.
The fluorescence can then be measured to determine the amount and type of cells present in a sample.
Up to thousands of particles per second can be analysed as they pass through the liquid stream.
A flow cytometer is made up of five main parts, as follows:
- A flow cell, which is the liquid stream or sheath fluid that arranges the cells in a sample so that they flow past the laser in single file.
- A measuring system, which is often a measure of conductivity and optical systems such as mercury or xenon lamps. Other examples include high-power water-cooled lasers such as argon, krypton or dye lasers. Low-power air-cooled lasers include argon (488 nm), red-HeNe (633 nm) or green-HeNe, HeCd (UV). Blue, green, red or violet diodelasers may also be used.
- An Analogue-to-Digital Conversion (ADC) system and a detector are also needed to generate FSC and SSC and also convert fluorescence signals from light to electrical signals that a computer can process.
- An amplifier, which may be either linear or logarithmic
- A computer for signal analysis
The process of flow cytometry involves the following:
- The term used to describe the collection of data form samples using flow cytometry is “acquisition.” This is achieved using a computer and software to ensure parameters such as voltage and compensation are correct for each sample.
- Modern instruments have several lasers and detectors as this enables multiple antibody labelling and accurate identification of target populations.
Reviewed by Sally Robertson, BSc