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Antibodies for Flow Cytometry

Flow cytometry is a technique for rapidly analyzing cells and particles as they stream in single file past one or more lasers. By recording light scatter, which corresponds to cell size and granularity, and the signals emitted by fluorescently-labeled antibodies for specific markers, flow cytometry allows for studying different cell types within a mixed population. The high-throughput capabilities of flow cytometry make it well suited to applications such as immunophenotyping, cell proliferation assays, and biomarker detection. In addition, cells of interest can be isolated for further downstream studies using fluorescence-activated cell sorting (FACS), a technique that is based on the same principles as flow cytometry.

Antibodies are fundamental to flow cytometry, enabling the specific detection of cell markers and other analytes. Fluorescently-labeled primary and secondary antibodies are respectively used for direct and indirect detection, while isotype control antibodies serve to reveal any non-specific background staining associated with primary antibodies. When selecting antibodies for flow cytometry, it is critical to identify well-validated products. The choice of fluorescent label is another important consideration, especially when building larger staining panels. We offer a broad range of primary antibodies that are validated for flow cytometry, along with matched isotype controls, as well as an extensive selection of fluorescently-labeled secondary antibodies.

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Frequently Asked Questions

Start by searching for antibodies that recognize your target of interest. Next, filter your search results to identify products that are validated for flow cytometry and have the desired species reactivity. Other factors to consider include the antibody host species, isotype, and antibody format (e.g., whole antibody or F(ab’)2 fragment), as well as which fluorescent labels are available. When selecting an antibody label, look for fluorophores that are compatible with your flow cytometer’s lasers and detectors, as well as with any other fluorophores being used in the same experiment.

Because monoclonal antibodies recognize just a single epitope, they are highly specific and present little risk of cross-reactivity. These features of monoclonal antibodies are critically important to avoid non-specific staining - a common challenge for flow cytometry, especially when detecting multiple targets. Monoclonal antibodies also benefit from exceptional lot-to-lot consistency and are available in unlimited supply, ensuring their reproducible long-term performance.

Yes, you can use polyclonal antibodies for flow cytometry. In some cases, they can improve sensitivity by providing signal amplification as they recognize multiple epitopes. However, keep in mind that polyclonal antibodies have a higher associated risk of cross-reactivity than monoclonal antibodies. Ways of addressing this include antigen-specific purification and cross-adsorption against closely related targets. In addition, each different lot of a polyclonal antibody will have a finite supply, making rigorous validation essential to ensure consistent performance.

Antibody validation should provide solid evidence that the antibody is specific for its target. Although the validation strategy will vary depending on the antigen, it is crucial to confirm that the antibody reacts as expected with endogenous samples. It is also recommended to compare the performance of the antibody with that of another antibody for the same target, ideally across multiple iterations of the same experiment, and to correlate the flow cytometry data with data from other applications. The inclusion of appropriate controls is essential to ensure the correct interpretation of results.

Related Product Categories

Flow Cytometry Reagents

Maximize fluorescence signal intensity and minimize non-specific binding in your flow cytometry assays

Isotype Controls

Move quickly and keep your assay development high quality and on track with our wide selection of isotype controls.

Primary Antibodies

Find a high-quality primary antibody from our fast-shipping selection, all produced to our exacting standards in our own facility.

Secondary Antibodies & Reagents

With a wide range of ready-to-ship secondary antibodies that are second to none, SouthernBiotech will keep your assays running and projects on schedule.