Immunomagnetic Cell Separation: Positive vs. Negative Selection
Whether to use a positive selection or negative selection approach is one of the most commonly asked questions regarding immunomagnetic cell separation. The right approach depends on your downstream application, required purity, and whether label-free cells are needed. ·¡²¹²õ²â³§±ð±èâ„¢ makes the choice simple, with clearly categorized kits and guidance to help you select the best option, while delivering reliable results through a straightforward workflow.
Why Use ·¡²¹²õ²â³§±ð±èâ„¢ to Isolate Cells?
- Isolate cells in as little as 8 minutes with a simple pour.
- Achieve up to 99% cell purities with high recoveries.
- Obtain viable, functional cells without the need for columns and washes.
- Isolate cells from virtually any sample source, including whole blood and leukopaks.
- Widely used in published research for over 20 years, supporting diverse downstream applications.
What Is the Difference Between Positive and Negative Selection?
Positive selection immunomagnetic cell separation methods directly label a desired cell type with an antibody that targets a specific cell surface protein. The antibody is linked to a magnetic particle, allowing the desired cells to be retained in the final isolated fraction after incubation of the sample in a magnetic field. In positive selection:
- Isolated cells are highly purified
- Isolated cells are usually bound by antibodies and magnetic particles
- Antibody cocktail targets a unique surface marker on the desired cells
- Additional cell populations can be isolated from the negative fraction
Negative selection immunomagnetic cell separation methods label unwanted cell types for removal with antibodies that target specific cell surface proteins. The antibodies are linked to magnetic particles, allowing the labeled, unwanted cells to be depleted from the isolated fraction by incubating the sample in a magnetic field. Since the desired cells are not specifically targeted by antibodies or ligands, they remain untouched and unbound by particles. In negative selection:
- Isolated cells can be highly purified or enriched
- Isolated cells are not bound by magnetic particles
- Antibody cocktail targets unwanted cells while leaving desired cells unlabeled
- Protocols are fast and easy with minimal sample manipulation
Figure 1. Comparison of Positive Selection and Negative Selection
How to Choose a Cell Selection Method
The choice between positive and negative selection methods often depends on your downstream application. For most applications, labeling cells with antibodies and ·¡²¹²õ²â³§±ð±èâ„¢ magnetic particles during positive selection does not interfere with downstream assays. However, you should always carefully consider whether your specific research application requires unlabeled cells, which may be important when:
- Antibody binding to a cell surface antigen is known to cause unwanted intracellular signaling
- Antibodies or magnetic particles bound to cell surface proteins affect downstream use of the cells
When in doubt, contact us to see whether we have data or recommendations for your specific application. Below are examples of common applications of immunomagnetic cell separation and the selection method we recommend.
Use Negative Selection for Quick Pre-Enrichment of Target Cells Prior to FACS
Isolating rare cell types by fluorescence-activated cell sorting (FACS) can require lengthy sort times. Researchers can substantially reduce cell sorting time by using immunomagnetic cell separation to pre-enrich their target cells. In this context, quick protocols and high recovery are essential to save time and maximize target cell yield resulting from the flow sorting procedure. Negative selection is typically an ideal approach because it is quick, has high recovery, and leaves the target cells unlabeled. This allows researchers to use any fluorophore-tagged antibodies against specific cell surface markers for downstream FACS analysis.
Pre-enrichment can be performed upstream of FACS for a wide variety of specific cell subsets, including antigen-specific cells, subsets of T helper cells, B cells, and innate lymphoid cells (ILCs).
View our fast negative selection kits ideal for pre-enrichment:
Use Indirect Positive Selection to Isolate Uncommon Cell Types
When isolating uncommon cells for which no specific, commercially available cell separation kits exist, indirect positive selection may be used. This method provides the flexibility of using your own primary antibodies to label the desired cells. Immunomagnetic cell separation can then be achieved by using commonly available secondary antibodies to attach magnetic particles to the target cells labeled by the primary antibodies. With this approach, almost any cell type can be isolated.
Conveniently, some cell separation providers also offer custom cell isolation kits that can be specifically designed for a particular starting cell source, species, and virtually any desired cell type or target cell surface antigen(s). For example, custom ·¡²¹²õ²â³§±ð±èâ„¢ kits allow you to isolate or enrich any cell type of interest.
Combine Selection Methods to Isolate Complex or Multiple Cell Types
Complex cell types may require a combination of negative and positive selection for successful purification. For example, isolating CD4+CD127lowCD25+ regulatory T cells (Tregs) can be challenging due to the need to select cells based on three different cell surface markers. Combining negative and positive selection strategies makes this possible. See the diagram below outlining the protocol for our ·¡²¹²õ²â³§±ð±èâ„¢ Human CD4+CD127lowCD25+ Regulatory T Cell Isolation Kit.
Figure 2. Protocol for the ·¡²¹²õ²â³§±ð±èâ„¢ Human CD4+CD127lowCD25+ Regulatory T Cell Isolation Kit
You can also isolate multiple cell types from a single sample through a sequential separation procedure. This approach may be particularly useful when your sample volume is limited and you do not wish to divide the sample.
Still Not Sure Which Method to Use?
Compare our ·¡²¹²õ²â³§±ð±èâ„¢ positive and negative selection kits to decide for yourself:
Get a Free ·¡²¹²õ²â³§±ð±èâ„¢ Sample
Curious about how ·¡²¹²õ²â³§±ð±èâ„¢ can simplify your research? Experience how easy it is by requesting a sample. Say goodbye to complicated protocols and time-consuming, column-based cell isolations. Make the easy choice and move your research forward faster.
Fully Automated Immunomagnetic Cell Separation with RoboSepâ„¢
Free up hands-on time and standardize your ·¡²¹²õ²â³§±ð±èâ„¢ cell isolations with RoboSepâ„¢-S or RoboSep-16 automated cell separation instruments.
On-Demand Immune Cell Isolation Courses
Join our free, self-paced training to learn how to effectively process human or mouse samples, and isolate highly pure target cells with ·¡²¹²õ²â³§±ð±èâ„¢.
Human Immune Cell Isolation Course
Learn how to effectively process human samples and isolate highly pure target cells with ·¡²¹²õ²â³§±ð±èâ„¢ in this free self-paced course.
Mouse Immune Cell Isolation Course
Learn how to isolate mouse immune cells quickly and easily from various sample sources with ·¡²¹²õ²â³§±ð±èâ„¢ and advance your immunology research.
Explore these Resources
