B7-H4 Monoclonal Antibody (188), PE, eBioscience™

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	cells were incubated on ice for 30min with appropriate antibodies or isotype controls, washed twice with FACS buffer (PBS containing 0.1% NaN3 and 5% fetal bovine serum), and resuspended in 0.5 ml 1% formalin/PBS. To analyze intracellular B7-H4, cells were pre-treated with Fix and Perm cell permeabilization reagents (Caltag Laboratories) according to the manufacturer's instructions.	All analyses were performed on a FACS calibur system (Becton Dickinson Immunocytometry Systems).

Protocol tips
cells were incubated on ice for 30min with appropriate antibodies or isotype controls, washed twice with FACS buffer (PBS containing 0.1% NaN3 and 5% fetal bovine serum), and resuspended in 0.5 ml 1% formalin/PBS. To analyze intracellular B7-H4, cells were pre-treated with Fix and Perm cell permeabilization reagents (Caltag Laboratories) according to the manufacturer's instructions.
Downstream tips
All analyses were performed on a FACS calibur system (Becton Dickinson Immunocytometry Systems).

Publication protocol

To detect surface B7-H4, CD11b or CD133 expression, cells were incubated on ice for 30min with appropriate antibodies or isotype controls (PE-B7-H4, PE-mouse IgG1, APC-CD11b, APC-CD133, APC-mouse IgG1, all from eBioscience, 1:5), washed twice with FACS buffer (PBS containing 0.1% NaN3 and 5% fetal bovine serum), and resuspended in 0.5 ml 1% formalin/PBS. To analyze intracellular B7-H4, cells were pre-treated with Fix and Perm cell permeabilization reagents (Caltag Laboratories) according to the manufacturer's instructions. Assays of immune function, cell proliferation, cell cycle, apoptosis, cytokines and cytotoxicity were done under standard methods as previously described. All analyses were performed on a FACS calibur system (Becton Dickinson Immunocytometry Systems).

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Manufacturer protocol

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