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Flow cytometry (FC) is a technique used to detect and measure the physical and chemical characteristics of a population of cells or particles. [1] [2] [3] [4]In this process, a sample containing cells or particles is suspended in a fluid and injected into the flow cytometer instrument.
Cell cycle analysis by DNA content measurement is a method that most frequently employs flow cytometry to distinguish cells in different phases of the cell cycle.Before analysis, the cells are usually permeabilised and treated with a fluorescent dye that stains DNA quantitatively, such as propidium iodide (PI) or 4,6-diamidino-2-phenylindole (DAPI).
Fluorescein isothiocyanate (FITC) is a derivative of fluorescein used in wide-ranging applications [1] [2] including flow cytometry.First described in 1942, [3] FITC is the original fluorescein molecule functionalized with an isothiocyanate reactive group (−N=C=S), replacing a hydrogen atom on the bottom ring of the structure.
The tetramers are labeled with a fluorophore, allowing tetramer-bound T-cells to be analyzed with flow cytometry. [4] Quantification and sorting of T-cells by flow cytometry enables researchers to investigate immune response to viral infection and vaccine administration as well as functionality of antigen-specific T-cells. [5]
Flow cytometry is by far the most sophisticated and expensive method for cell counting. In a flow cytometer the cells flow in a narrow stream in front of a laser beam. The beam hits them one by one, and a light detector picks up the light that is reflected from the cells.
A wide (hundreds of micrometers in diameter) tube made of glass or plastic is used, through which a "wall" of fluid called the sheath flow is pumped. The sample is injected into the middle of the sheath flow. If the two fluids differ enough in their velocity or density, they do not mix: they form a two-layer stable flow. [2]
Cytometry is the measurement of number and characteristics of cells. Variables that can be measured by cytometric methods include cell size , cell count , cell morphology (shape and structure), cell cycle phase, DNA content, and the existence or absence of specific proteins on the cell surface or in the cytoplasm . [ 1 ]
Droplet sorting in microfluidics is an important technique, allowing for discrimination based on factors ranging from droplet size to chemicals labeled with fluorescent tags within the droplet, stemming off of the work done to sort cells in Flow Cytometry.
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