Anti-Hes1 antibody [EPR4226] (ab108937) at 1/1000 dilution (purified) + SH-SY5Y cell lysate at 10 µg

Secondary
HRP goat anti-rabbit (H+L) at 1/1000 dilution

Predicted band size: 30 kDa
Observed band size: 30 kDa

Blocking buffer: 5% NFDM/TBST

Dilution buffer: 5% NFDM/TBST

Primary ab concentration dilution: 1:200, (0.5ug/ml), Secondary ab: ImmunoHistoprobe (Ready to use) HRP Polymer for Rabbit IgG, Secondary ab concentration: Prediluted, Tissue: Human brain, Fixative: Paraffin-embedded sections, Counter stain: Hematoxylin Antigen retrieval: Perform heat mediated antigen retrieval using Tris/EDTA Buffer, PH9

Immunofluorescent staining of SH-SY5Y cells (fixed with 4% PFA and permeablized with TritonX 100) with purified ab108937 at a dilution of 1/100. An Alexa Fluor^® 555 goat anti-rabbit antibody was used as the secondary at a dilution of 1/200. The panel on the right shows the DAPI counter-staining.

Overlay histogram showing SH-SY5Y cells stained with unpurified ab108937 (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. The cells were then incubated with the antibody (ab108937, 1/100 dilution) for 30 min at 22ºC. The secondary antibody used was DyLight^® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1µg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed.

All lanes : Anti-Hes1 antibody [EPR4226] (ab108937) at 1/1000 dilution (unpurified)

Lane 1 : Fetal brain lysate
Lane 2 : PC12 cell lysate
Lane 3 : SH-SY5Y cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : HRP-labelled goat anti-rabbit at 1/2000 dilution

Predicted band size: 30 kDa

Immunohistochemical staining of Hes1 in paraffin-embedded human placenta tissue with unpurified ab108937 at 1/250 dilution.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Immunohistochemical staining of paraffin embedded human brain with unpurified ab108937 at a working dilution of 1 in 90. The secondary antibody used is a HRP polymer for rabbit IgG. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0.

Immunofluorescent staining of SH-SY5Y cells (fixed with 4% PFA and permeablized with TritonX 100) with unpurified ab108937 at a dilution of 1/40. An Alexa Fluor^® 555 goat anti-rabbit antibody was used as the secondary at a dilution of 1/200. The panel on the right shows the DAPI counter-staining.