Remove supernatant by careful aspiration. Analyze in a flow cytometer according to manufacturer's instructions. Proper color compensation is important for unbiased data interpretation. Cell samples stained with the corresponding single reagents of the pair may be used as controls for adjusting compensation. It is advisable to run the appropriate negative controls.
Negative controls establish background fluorescence and non-specific binding of the antibodies.
The degree of autofluorescence or negative control reagent fluorescence will vary with the type of cells under study and the sensitivity of the instrument used. Beverley, P., et al., Eur. Borst, J., et al., J. Biol. Callard, R., et al., Clin. Fischer, A., et al.