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Star Republic: Guide for Biologists

Membrane immunostaining

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Membrane proteins can be stained with antibody without fixing cells. This protocol stains cells in suspension for FACS analysis.

1. Add 25-50 Ál diluted antibody (in PBS) to 5x105 cells (25 Ál).

2. Incubate on ice for 30 minutes.

3. Wash twice with PBS/1% BSA/0.02% NaN3.

4. Add 50 Ál of an appropriate FITC-labeled secondary antibody.

5. Incubate on ice for 30 minutes.

6. Wash twice with PBS/1% BSA/0.02% NaN3.

7. Resuspend the cells in 100 Ál PBS/1% BSA/0.02% NaN3 and analyze on a fluorescence activated cell sorter (FACS).