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Flow Cytometry Animation

A brief introduction to flow cytometry from Cell Signaling Technology (CST). Visit https://www.cellsignal.com/flowcytometry for more information on validated products and optimized protocols for flow cytometry. Transcript: Cell processes are controlled by complex and dynamic intracellular signaling networks. Surface receptors pass information into the cell by modifying cytoplasmic proteins. These modifications include phosphorylation,. methylation, and acetylation, which control a broad range of cellular functions, including growth, development, and apoptosis. Disruption of signaling networks often leads to disease. To understand signaling processes and normal and pathological conditions, identification and quantification of signaling proteins is necessary. Flow cytometry is ideal for such a task. Flow cytometry is a laser-based technology widely used to phenotype cells using antibodies directed against surface markers. It can also allow simultaneous analysis of multiple intracellular targets using antibodies that detect and signaling proteins, their post-translational modifications, epigenetic modifications, and downstream transcription factors. The first step of intracellular flow cytometry is fixation of the cells using formaldehyde, which immobilizes proteins and preserves transient signaling events. Cells are then permeabilized with methanol or detergent, allowing antibodies to access to the intracellular space. Optimal fixation and permeabilization conditions depend on the location and biochemistry of the target epitopes. It should be carefully considered to ensure successful antigen detection, especially when staining surface and intracellular proteins simultaneously. Since multiple signaling proteins and surface antigens can be identified using fluorescently distinct antibodies, flow cytometry can readily characterize signaling profiles at the single-cell level within phenotypically distinct populations. This allows the characterization of the relationship between intracellular targets and enables detection of signaling events from rare-cell populations within a heterogeneous sample. Multiplex analysis of these protein targets saves time and resources while producing data-rich results, making flow cytometry an invaluable tool to unravel the complexities of cell signaling.