ab1793 staining TNFα in RAW 264.7 cells treated with (R,S)-rolipram (ab120029), by ICC/IF. Decrease in TNFα expression correlates with increased concentration of (R,S)-rolipram, as described in literature.
The cells were incubated at 37°C for 24h in media containing different concentrations of ab120029 ((R,S)-rolipram) in DMSO, fixed with 100% methanol for 5 minutes at -20°C and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab1793 (5 µg/ml) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight 488 goat anti-mouse polyclonal antibody (ab96879) at 1/250 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI and are shown in blue.

Overlay histogram showing RAW 264.7 cells stained with ab1793 (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 (ab1793, 1/10 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a decreased signal in RAW 264.7 cells fixed with 4% paraformaldehyde (10 min) used under the same conditions.

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

ab1793 staining TNFα in RAW 264.7 cells treated with (R)-(-)-rolipram (ab120031), by ICC/IF. Decrease in TNFα expression correlates with increased concentration of (R)-(-)-rolipram , as described in literature.
The cells were incubated at 37°C for 24h in media containing different concentrations of ab120031 ((R)-(-)--rolipram) in DMSO, fixed with 100% methanol for 5 minutes at -20°C and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab1793 (5 µg/ml) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight 488 goat anti-mouse polyclonal antibody (ab96879) at 1/250 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI and are shown in blue.

ab1793 staining TNFα in RAW 264.7 cells treated with (S)-(+)-rolipram (ab120030), by ICC/IF. Decrease in TNFα expression correlates with increased concentration of (S)-(+)-rolipram , as described in literature.
The cells were incubated at 37°C for 24h in media containing different concentrations of ab120030 ((S)-(+)-rolipram) in DMSO, fixed with 100% methanol for 5 minutes at -20°C and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab1793(5 µg/ml) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight 488 goat anti-mouse polyclonal antibody (ab96879) at 1/250 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI and are shown in blue.