Anti-HCN1 antibody [S70-28] (ab84816) + rat brain lysate

Overlay histogram showing SH-SY5Y cells stained with ab84816 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab84816, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. 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 positive result in SH-SY5Y cells fixed with 4% paraformaldehyde (10 min)/permeabilized in 0.1% PBS-Tween for 20 min used under the same conditions.

Immunohistochemical analysis of murine cochlear tissue, staining HCN1 (red) with ab84816. Tissue was permeabilized with 0.3% Triton X-100 before being blocked with goat serum. Sections were incubated with primary antibody (1/100) overnight at 4°C. An AlexaFluor®568-conjugated goat anti-mouse IgG (1/2000) was used as the secondary antibody.