Usually the company where antibodies come from is supposed to provide a detailed protocol for you. Have you tried contacting a representative?

dear alexandra , yess i have contacted a representitive but she gived me a protocol in general . and so complicated and not detailed , so i need a protocol detailed and clear .

thanks

That is strange they do not give you more advice. The problem is each antibody might have a specific protocol, although in general flow cytometry staining is pretty simple - you incubate your cells in BSA, then just apply your primary to cell suspension, resuspend, incubate, centrifuge, take out supernatant, resuspend in your secondary and take it to FACS. Since it's a surface marker, you do not need to penetrate cells. And if there are any specific varieties to the protocol - they depend primarily on the company where the antibody comes from. I would keep asking them for more detail. Maybe you have specific questions?

These links may help you

http://www.abcam.com/protocols/indirect-flow-cytometry-protocol

http://www.abcam.com/protocols/direct-flow-cytometry-protocol

https://www.rndsystems.com/resources/protocols/flow-cytometry-protocol-analysis-cell-viability-using-propidium-iodide

https://www.bio-rad-antibodies.com/flow-cytometry-protocols-procedures.html

https://www.thermofisher.com/my/en/home/references/protocols/cell-and-tissue-analysis/flow-cytometry-protocol.html

https://www.rndsystems.com/resources/protocols/flow-cytometry-protocols

https://www.biomol.de/dateien/protocols/NSJBio/NSJBio--Flow-Cytometry-Protocol.pdf

http://www.sigmaaldrich.com/life-science/cell-biology/cancer-research/learning-center/cancer-research-protocols/flow-cytometry.html

https://facs.stanford.edu/sites/default/files/documents/scheduler/flowcytometry-intro.pdf

http://www.mitosciences.com/PDF/flow-cytometry-protocol.pdf

Is it compulsory to use fresh blood for flow cytometry?