CST BLOG: Lab Expectations

The official blog of Cell Signaling Technology® (CST) where we discuss what to expect from your time at the bench, share tips, tricks, and information.

Need cell type and function? Use flow cytometry!

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Need cell type and function? Use flow cytometry!

What is flow cytometry and how is it used?

Flow cytometry enables you to save time and analyze many characteristics of your cells in one experiment, using classic principles of antibody detection.


By selecting and validating a suitable panel of antibodies, you can develop a flow cytometry experiment to detect intracellular signaling states, and identity of thousands of cells all from one cell preparation. Importantly, because of flow cytometry provides a quantitative readout, it can more accurately identify, enumerate, and subdivide populations of cells exhibiting a range of defined characteristics in just a few hours. This makes flow cytometry the ideal tool for the generation of robust datasets in a very short timeline.

While flow cytometry has many uses in research and drug development, it is particularly helpful when assessing tumor responses to various anti-cancer therapies. Tumor samples are composed of heterogeneous mixtures of cells, and analysis of drug-driven changes requires the ability to capture and quantify unique responses from all represented cell types.

Flow cytometry is also particularly helpful in the field of immunology where the identification of various immune cells and characterization of immune-related signaling states are essential.

Flow cytometry can characterize biological processes that are indicative of cellular function/dysfunction in unique cell types. This analysis can be performed on a multitude of signaling states in a heterogeneous sample.

Flow cytometry is capable of analyzing a multitude of signaling states in cells within a heterogenous sample, to better identify and characterize unique biological processes that are indicative of cellular function or dysfunction or those indicative of a unique cell type.

In addition, one other major benefit of flow cytometry is that, unlike other protein analysis methods that require a large number of cells for protein isolation and analysis, flow cytometry is much better suited for the assessment of patient-derived tissue samples or primary cells that exist in very limited quantities.

All of these unique benefits make flow cytometry an ideal tool for time-efficient, cost-effective immunophenotyping of cells in various research areas.

Important considerations that can impact a flow cytometry experiment

For the success of a flow cytometry experiment, researchers must first select appropriate primary antibodies that have been validated for binding-affinity and specificity to the chosen target.  This is important if the user aims to discriminate between unique cell types or states based on small differences in expressed proteins. In addition, because flow cytometry relies on fluorescent detection, investigators must also carefully choose fluorescent conjugates that minimize spectral overlap. This is particularly important when multiple parameters are being analyzed in a single experiment. Fluorophores can be conjugated directly to the primary antibody or indirectly to a secondary antibody that is matched by species and immunoglobulin isotype to the primary antibody. In the case of indirect detection, varying species and isotypes of the primary antibody can greatly increase the number of targets simultaneously detected in each experiment. Thus, careful planning and selection of these three parameters would ensure proper detection of multiple unique antigens in one experiment, thereby increasing data quality and robustness. 

As a leading producer of high-quality antibodies, CST has hundreds of ready-to-use or custom-made flow-validated antibody conjugates in a variety of fluorophores that are designed to reproducibly detect essential cellular characteristics. CST also has comprehensive technical support to help design and carry out flow cytometry experiments.

Learn more about flow cytometry.

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