Flow cytometric analysis for floating cells:
We usually use Fisher tubes or equivalents as reaction tubes for all step described below.
1) Wash the cells 3 times with washing buffer [PBS containing 2% fetal calf serum (FCS) and 0.1% NaN3].
2) Resuspend the cells with washing buffer (5x10^6cells/mL).
3) Add 50 uL of the cell suspension into each tube, and centrifuge at 500 x g for 1 minute at room temperature
(20~25 degrees Celsius). Remove supernatant by careful aspiration.
4) Add 10 uL of normal goat serum containing 1 mg/mL normal human IgG and 0.1% NaN3 to the cell pellet after tapping. Mix well and incubate for 10 minutes at room temperature.
5) Add 40 uL of the primary antibody at the concentration of as suggest in the APPLICATIONS diluted in the washing buffer. Mix well and incubate for 30 minutes at
6) Add 1 mL of the washing buffer followed by centrifugation at 500 x g for 1 minute at room temperature. Remove supernatant by careful aspiration.
7) Resuspend the cells with 500 uL of the washing buffer and analyze by a flow cytometer.