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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 vialswith 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
  • ThreeBD 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):

  1. Verify origin of your antibody (Hamster, Rat or Mouse) to specify which beads should be used
  2. Mix Beads well by shaking the bottle
  3. Pre-dilute anti-Ig Beads using 400 ul of PBS for 1 drop of Beads
  4. Dispense 100 ul of pre-diluted Beads per single color control sample
  5. Add 5 ul of pre-diluted primary Ab to the Beads
  6. Incubate 20 min at RT, mix occasionally
  7. If applicable, add 5 ul of pre-diluted secondary Ab with Fluorochrome
  8. Incubate 20 min at RT, mix occasionally
  9. Add 50 um of pre-diluted uncoated Beads to each sample
  10. Make separate tube with uncoated beads for Negative Control
  11. Use immediately for Multi-color compensation or store at +4oC for up to 2 weeks