Flow cytometry is a semi-quantitative technique in biotechnology used for cell counting, cell sorting, biomarker detection, and protein engineering. The technique is commonly used to monitor the immune response by measuring the frequency and functionality of different immune cells. The instrument used is known as flow cytometer. The cells stained with specific fluorochrome-conjugated antibodies are passed through the flow cytometer to analyse the frequency and other properties of the cells.
Here are the various uses of flow cytometry in preclinical studies:
Flow cytometers can measure a single cell suspension ranging from the immune cells of the blood to the tissue culture cells and antibodies that can bind immune molecules like cytokines. Simultaneously, they can also measure multiple cell types and differentiate them based on size and granularity.
The Staining Panel
Flow cytometry utilises fluorochrome-conjugated antibodies. Several fluorochromes and fluorochrome combinations have been identified that attach to the specific antibodies and emit photons at discrete wavelengths. Thus, each antibody can be utilised to recognise a specific molecule on the cell’s surface or interior, and thus detect a specific cell population. There are different fluorochrome-conjugated antibodies for immune cell markers in different species such as humans, mouse, primates, etc.
How do Flow Cytometers Work?
Flow cytometers use lasers to measure the cells labelled with fluorochrome antibodies. They direct lasers at specific wavelengths at a stream of individual cells stained with fluorescent antibodies. These cells travel through the flow cytometer in a liquid called sheath fluid. The flow cytometers emit multiple lasers at different wavelengths to measure a variety of fluorochromes. The laser excites fluorochrome-conjugated antibodies bound to the cells. This results in the movement of electrons from the fluorescent molecules to a higher energy state for a short period after which they return to their ground state. The laser also results in the emission of photons at a higher wavelength from the fluorochrome-conjugated antibodies. These photons pass through the filters to reach an appropriate detector. They are further converted into a voltage and all this event is captured by computer software.
The software analyses these events and the data is represented to show different cell parameters like size, shape, granularity, and frequency of antibody-labelled cells. There are different types of flow cytometers with different lasers and emission detectors, but each cytometer has a unique configuration of excitation and emission parameters. And this must be considered while selecting the antibodies for a staining panel.
Flow cytometry is used to perform different procedures including:
- Cell counting
- Cell sorting
- Detection of biomarkers
- Protein engineering
By adding flow cytometry to your preclinical research program, you can really propel it forward faster, save time in analyzing the samples and get to work with flow cytometry professionals. Working with flow cytometry professionals from specialized contract research organisations will ensure you get the expertise required for your preclinical studies.