Bead Compensation Controls
Reason to use BD FACSComp Bead as Single Stain Controls:
- Cell numbers are limiting
- Antigen is dimly expressed
- Antibody has low affinity to receptor
- Positive population is rare
- Use tandem dyes
BD FACSComp Bead — Kits.
Details and ordering information:
- Each Kit contains two vials with bead polystyrene particles; 6ml each; $180.00
- Beads uncoated (FBS) that have no binding capacity; used for negative population
- Beads coated with Goat anti-Ig kappa which bind light chain-bearing immunoglobulin; used for positive population.
Be sure to use same Fluorochrome-conjugated monoclonal antibodies as actually used for cell staining in your experiment
- Three BD FACSComp Bead Kits available with the coated beads of different specificity:
- Anti-Mouse Ig, kappa (Cat#552843, BD Biosciences)
- Anti-Rat Ig, kappa (Cat#552844, BD Biosciences)
- Anti-Rat/Hamster Ig, kappa (Cat#552845, BD Biosciences)
BD FACSComp Beads labeling procedure
(implemented and recommended by FCRC):
- Verify origin of your antibody (Hamster, Rat or Mouse) to specify which beads should be used
- Mix Beads well by shaking the bottle
- Pre-dilute anti-Ig Beads using 400 ul of PBS for 1 drop of Beads
- Dispense 100 ul of pre-diluted Beads per single color control sample
- Add 5 ul of pre-diluted primary Ab to the Beads
- Incubate 20 min at RT, mix occasionally
- If applicable, add 5 ul of pre-diluted secondary Ab with Fluorochrome
- Incubate 20 min at RT, mix occasionally
- Add 50 um of pre-diluted uncoated Beads to each sample
- Make separate tube with uncoated beads for Negative Control
- Use immediately for Multi-color compensation or store at +4oC for up to 2 weeks
