BD, 569991, BD™ SpectraComp™ Unmixing and Compensation Particles

Reactivity: Mouse, Rat, Hamster (Tested in Development)
Application: Flow cytometry (Routinely Tested)
Vol. Per Test: 1 drop/test
Storage Buffer: Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
Regulatory Status: RUO
Preparation And Storage: Preparation And Storage Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.
Recommended Assay Procedures: Recommended Assay Procedures The BD™ SpectraComp™ Unmixing and Compensation Particles have been tested with mouse, rat, and hamster Ig antibodies conjugated to various fluorochromes. See the specific instructions below on the use of the BD™ SpectraComp™ Unmixing and Compensation Particles. 1. Vortex BD™ SpectraComp™ Unmixing and Compensation Particles thoroughly before use. 2. Add 1 drop (approximately 50 µl) of BD™ SpectraComp™ Unmixing and Compensation Particles into the bottom of 12 × 75 mm tubes or 96-well plate wells for each fluorochrome used in the experiment. It is recommended to include one tube for unstained control. 3. Add antibody reagent to the mixture and vortex.  Incubate at room temperature for 15-30 minutes, protected from light. Notes: • It is recommended to predetermine the appropriate amount of antibody reagent that works best for the application. • Treat the BD™ SpectraComp™ Unmixing and Compensation Particles with the same protocol used for the cell sample, eg, if the testing protocol uses BD Horizon™ Brilliant Stain Buffer or permeabilized and fixed cells, the particles should also be treated with the reagents and protocol. 4. Add 2 mL of staining buffer [eg, BD Pharmingen™ Stain (FBS), Cat. No. 554656 or BD Pharmingen™ Stain (BSA), Cat. No. 554657] to the tubes or required volume to the plate wells. 5. Wash the particles by centrifuging the tubes or plate for 5 minutes at 500 × g and immediately aspirate the supernatants to minimize particle loss, being careful not to disturb the pellet. Wash two times by repeating steps 4 and 5 leaving approximately 50 µL of supernatant in the tube after each aspiration. Note: • For best signal-to-noise results, use a vacuum aspirator and aspirate off the supernatant using a fine pipette tip. 6. Resuspend the pellet by vortexing and by then adding 0.3-0.5 mL staining buffer. 7. Acquire the BD™ SpectraComp™ Unmixing and Compensation Particles using the same Forward and Side Light Scatter parameter (FSC-A and SSC-A) instrument settings used for analyzing cells. It is recommended to acquire the data immediately after staining. 8. Gate on the singlet bead population based on FSC (forward-light scatter) and SSC (side-light scatter) characteristics. 9. For compensation or spectral unmixing use the appropriate protocol for your cytometer system. 10. Proceed to acquire the cell samples for the experiment. It is recommended to test the BD™ SpectraComp™ Unmixing and Compensation Particles to confirm accuracy of compensation or spectral unmixing.