In a perfect world, the fluorescence emission profile for each individual fluorophore would be a very intense, narrow peak, well separated from all other emission peaks. In reality, organic dyes and fluorescent proteins have broad emission peaks, and compensation must be employed (during or after data acquisition) to correctly assign fluorescence signal to each fluorophore. Our flow cytometry compensation beads are manufactured with low lot-to-lot variability and proven long-term stability to help ensure that you get consistent and reliable compensation in every experiment. They may be used to augment or replace your cell-based compensation controls, depending on your samples and application.
Flow Cytometry Compensation Beads
Flow cytometry compensation beads selection guide
|AbC Total Antibody Compensation Bead Kit||AbC Anti-Mouse Bead Kit||AbC Anti-Rat/Hamster Bead Kit||ArC Amine Reactive Compensation Bead Kit|
|Assay type||Immunophenotyping||Immunophenotyping||Immunophenotyping||Cell viability assay|
|Positive bead||Hamster, mouse, rabbit, and rat antibodies*||Mouse monoclonal antibodies*||Rat and hamster monoclonal antibodies*||Amine reactive dyes and LIVE/DEAD Fixable Dead Cell Stains|
|Negative bead||No binding capacity||No binding capacity||No binding capacity||No amine reactive capacity|
|Format||100 reactions||25 reactions||100 reactions||100 reactions||100 reactions||25 reactions|
|*AbC capture beads bind all isotypes.|
Compensation beads for immunophenotyping
AbC Compensation Bead Kits
- An alternative to using precious samples for setting flow cytometry compensation
- Highest reactivity to different subclasses of mouse, rat, and hamster immunoglobulin
- Fast and simple bead-based fl ow cytometry compensation
- Removal of inconsistencies due to variations in antigen expression
Invitrogen™ AbC™ bead kits provide a consistent, accurate, and simple-to-use technique for the setting of flow cytometry compensation when using:
- Fluorophore-conjugated mouse, rat, hamster, or rabbit antibodies (AbC Total Antibody Compensation Bead Kit) (Figure 1)
- Fluorophore-conjugated mouse antibodies (AbC Anti-Mouse Bead Kit) or
- Fluorophore-conjugated rat or hamster antibodies (AbC Anti-Rat/Hamster Bead Kit)
All three kits contain two types of specially modified polystyrene microspheres: 1) AbC capture beads (also called positive beads), which bind all isotypes of the specific immunoglobulin, and 2) negative beads, which have no antibody binding capacity. After incubation with a fluorophore conjugated primary antibody (mouse, rat, hamster, or rabbit, depending on the kit used), the two components provide distinct positive and negative populations of beads that can be used to set compensation. You can perform compensation with the same fluorophore labeled antibody used for cell staining. The consistent nature of bead scatter and high surface antibody-binding capacity allows you to more consistently and accurately set compensation for any combination of fluorophore-labeled primary antibodies. Both types of microspheres (the AbC capture beads and negative beads) have a diameter of approximately 6 μm (actual size for each lot is listed on the component vial). The bead suspensions are supplied in dropper vials for convenient sample application.
|Figure 1. Histograms showing staining of the Invitrogen™ AbC™ Total Antibody Compensation Bead Kit. The histograms show the signal separation of the positive capture beads from the negative beads after binding to mouse (top left), rat (top right), and hamster (bottom left) monoclonal antibodies, and rabbit (bottom right) mono- and polyclonal antibodies. Capture beads were labeled with an optimized amount of each PE antibody conjugate and analyzed on an Attune™ Acoustic Focusing Cytometer using 488 nm excitation and a 574/26 nm bandpass filter.|
Compensation beads for LIVE/DEAD Fixable Dead Cell Stains
ArC Amine Reactive Compensation Bead Kit
- Eliminates the hassle of heat-treating cells
- Optimized for all LIVE/DEAD Fixable Dead Cell Stain kits
- Fast and simple bead-based flow cytometry compensation
- An alternative to using precious samples for setting compensation
- Accurate and consistent results
The Invitrogen™ ArC™ Amine Reactive Compensation Bead Kit provides a consistent, accurate, and simple-to-use technique for the setting of flow cytometry compensation when using any of the Invitrogen™ LIVE/DEAD™ Fixable Dead Cell Stain Kits or when using any amine reactive dye. LIVE/DEAD Fixable Dead Cell Stain Kits (and amine reactive dyes) can be used to evaluate mammalian cell viability based on the fact that the dye reacts with cellular amines. The reactive dye can enter the cell via compromised membranes of necrotic cells and react with free amines in the interior and on the surface of the cell, resulting in intense fluorescent staining. In contrast, only the cell-surface amines of viable cells are available to react with the dye, resulting in relatively dim staining. The difference in fluorescence intensity between the live and dead cell populations is typically greater than 50-fold.
The ArC Amine Reactive Compensation Bead Kit includes two types of specially modified polystyrene microspheres to allow easy compensation of the LIVE/DEAD Fixable Dead Cell Stains: the ArC reactive beads (Component A), which bind any of the amine reactive dyes, and the ArC negative beads Component B), which have no reactivity. After incubation with any amine reactive dye, the two kit components provide distinct positive and negative populations of beads that can be used to set compensation (Figure 2). The ArC Amine Reactive Compensation Bead Kit can be combined with the AbC Total Antibody Compensation Bead Kit for use with fluorophore conjugated mouse antibodies, allowing even more consistent and accurate compensation for multicolor immunophenotyping experiments that also incorporate a LIVE/DEAD Fixable Dead Cell Stain.
Figure 2. Staining profile of the Invitrogen™ ArC™ Amine Reactive Compensation Bead Kit components with three Invitrogen™ LIVE/DEAD™ Fixable Dead Cell Stain Kits. (A) Invitrogen™ LIVE/DEAD™ Fixable Violet dye stained beads were analyzed with 405 nm excitation, emission was collected with a 450/50 nm bandpass filter. (B) Invitrogen™ LIVE/DEAD Fixable Green dye stained beads were analyzed with 488 nm excitation, emission was collected with a 525/50 nm bandpass filter. (C) Invitrogen™ LIVE/DEAD Fixable Far Red dye stained beads were analyzed using 633 nm excitation, emission was collected with a 660/20 nm bandpass filter.
For Research Use Only. Not for use in diagnostic procedures.