ab6640 stained Raw 264.7 cells. The cells were 100% methanol fixed for 5 minutes at room temperatureand then incubated in 1%BSA / 10% normal Goat serum / 0.3M glycine in 0.1% PBS-Tween for 1hour at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab6640 at 5µg/ml) overnight at +4°C. The secondary antibody (pseudo-colored green) was ab150165 Goat Anti-Rat IgG H&L (Alexa Fluor® 488) preadsorbed used at a 1/1000 dilution for 1 hour at room temperature. Alexa Fluor® 594 WGA was used to label plasma membranes (pseudo-colored red) at a 1/200 dilution for 1hour at room temperature. DAPI was used to stain the cell nuclei (pseudo-colored blue) at a concentration of 1.43µM for 1hour at room temperature.

Flow cytometry analysis of J774 cells labelling F4/80 (red) with ab6640 at a dilution of 1/10 followed by streptavidin FITC secondary at 1/100 dilution. The blue line shows J774 cells stained with Rat anti mouse isotype control. 

ab6640 staining F4/80 in mouse bone marrow cells by immunocytochemistry/ immunofluorescence. Cells were formaldehyde fixed and permeabilized in 0.2% Triton X-100 prior to blocking in 2% BSA for 30 minutes at 20°C. The primary antibody was diluted 1/200 and incubated with the sample for 9 hour at 4°C. Alexa fluor® 488 goat polyclonal to rat Ig, diluted 1/200, was used as the secondary antibody.

See Abreview

Flow cytometry analysis of peritoneal macrophages labelling F4/80 (red) with ab6640 at a dilution of 1/10. The blue line shows J774 cells stained with Rat anti mouse isotype control. 

Flow cytometry analysis of J774 cells labelling F4/80 (red) with ab6640 at a dilution of 1/50 followed by goat anti rat IgG FITC secondary antibody at 1/100 dilution. The blue line shows J774 cells stained with Rat anti mouse isotype control.