Detection of F4/80 in RAW cells. Red, black and blue line represent the isotype control, cells only and ab16911 at 10 μg/ml, respectively.

ab16911 staining F4/80 on macrophages in mouse liver tissue by Immunohistochemistry (Frozen sections).

ab16911 staining F4/80 in Mouse brain cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with acetone and blocked with 5% BSA for 1 hour at 20°C. Samples were incubated with primary antibody (1/250) for 16 hours at 4°C. An Alexa Fluor^®568-conjugated Goat anti-rat IgG polyclonal (1/1000) was used as the secondary antibody.

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Overlay histogram showing HeLa cells stained with ab16911 (red line). The cells were fixed with methanol (5 min) and incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab16911, 1/10 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rat IgG (Fc) (ab96971) at 1/250 dilution for 30 min at 22°C. Isotype control antibody (black line) was rat IgG2a [aRTK2758] (ab18450, 2µg/1x10⁶ cells cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a significantly decreased signal in HeLa cells fixed with 4% paraformaldehyde (10 min) used under the same conditions.

Please note that Abcam does not have data for use of this antibody on non-fixed cells. We welcome any customer feedback.

ab16911 staining mouse spleen tissue sections by immunohistochemistry (frozen sections). Sections were paraformaldehyde fixed without permeabilization and blocked in 1% serum for 10 minutes at 20°C. The primary antibody was used undiluted and incubated with sample for 16 hour at 20°C. A Biotin conjugated goat polyclonal to rat Ig, diluted 1/500 was used as the secondary antibody.

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ab16911 stained RAW246.7 cells. The cells were 100% methanol fixed for 5 minutes at -20°C and 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 (ab16911 at 1in50 dilution) overnight at +4°C. The secondary antibody (pseudo-colored green) was Goat Anti-Rat IgG H&L (Alexa Fluor® 488) preadsorbed (ab150165) used at a 1/1000 dilution for 1hour 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.