Overlay histogram showing human CD14+ monocytes stained with ab221248 (red line). Human peripheral blood mononuclear cells were incubated for 30min on ice in 1x PBS containing 10ug/ml human IgG and 10% normal goat serum to block FC receptors and non-specific protein-protein interactions. Then cells were surface stained for CD14 FITC (ab28061) for 30 min on ice. The cells were then permeabilised with 4.2% formaldehyde and 0.1% saponin for 20 min at 4°C. After cells were blocked with 1xPBS containing 0.1% saponin and 10% normal goat serum for 30 min on ice followed by the antibody (ab221248, 1x10⁶ cells in 100ul at 1ug/ml) for 30 min on ice.

Isotype control antibody (black line) Rabbit IgG (monoclonal) Allophycocyanin (ab232814) used at the same concentration and conditions as the primary antibody. CD14 FITC single stained cells (blue line) were also used as a control.

Acquisition of >5,000 events were collected using a 40 mW Red laser (640nm) and 670/14 bandpass filter. Events were gated on live single cell CD14+ positive monocytes.