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FLOW CYTOMETRY

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Flow cytometry is a technology that provides rapid multi-parametric analysis of single cells in solution.

Flow cytometry is a technology that provides rapid multi-parametric analysis of single cells in solution. It enables recognition, characterisation and subsequent sorting of cells of interest. Flow cytometry simultaneously measures multiple physical characteristics of single particles, usually cells, as they flow in a liquid stream past a light source, typically a laser. The properties measured include the particle’s relative size and the relative fluorescent intensity. The brightness of the fluorescence correlates to the amount of the measured property such as the amount of protein (eg number of receptors on a cell) or amount of DNA.

 

Fluorophores are the “colours” used to tag cellular properties of interest. Each fluorophore emits light of a specific wavelength when excited by the correct laser. This means that the fluorescently stained cells or particles can be detected individually, and this assists in the recognition, classification and quantification of different immune cell types (also called immunophenotyping). A specialised flow cytometer called a Fluorescence Activated Cell Sorter (FACS) subsequently allows for the isolation of particles of interested based on the selected criteria or immunophenotype.

 

In the Hugh Green Cytometry Centre we use the multiple lasers and detectors on our high-end full spectrum flow cytometers to allow us to label cells with more than 30 individual fluorophores conjugated to antibodies, for immunophenotyping and characterisation. The ability to simultaneously measure multiple parameters on a cell-by-cell basis is one of the most powerful aspects of analytical flow cytometry.  

 

Aside from immunophenotyping there are many other applications of flow cytometry, including intracellular antigen expression (cytokine/chemokine production), cell cycle analysis, measurement of DNA and RNA content, apoptosis, viability, rare event analysis (stem cells, side population, dendritic cells), cell counting (absolute counts), fluorescent protein expression, mitochondrial membrane potential, cellular kinetics (proliferation), FRET, bead assays (CBAs), bacteria analysis and cell sorting. From immune cells, nanoparticles, microalgae and bacteria to parasite eggs, if the sample can be reduced to individual particles ranging from 0.1µm-60µm we can analyse and sort them at speeds of >10,000 cells per second.

Our team ensure that the flow cytometers housed in the Hugh Green Cytometry Centre undergo daily quality control checks and are kept in optimal working order. We offer instrument support and troubleshooting tips, panel design, validation and implementation support, assistance with experiment optimisation such as sample preparation, troubleshooting and experimental design; data analysis, interpretation, troubleshooting and reporting.

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