IHC image of CD1d staining in human normal skin formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with EDTA based pH 9.0 solution (epitope retrieval solution 2) for 20 mins. The section was then incubated with ab11076, 5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

Overlay histogram showing MOLT4 cells stained with ab11076 (red line). The cells were fixed with 4% paraformaldehyde (10 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 (ab11076, 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. Unlabeled sample (blue line). Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

ab11076 staining CD1d in human skin tissue by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections).
Tissue was fixed with formaldehyde and an antigen retrieval step was performed using EDTA pH 8.0 for 20 minutes at 100°C. Samples were then incubated with ab11076 at a 1/200 dilution for 20 minutes at 25°C. The secondary used was an undiluted HRP conjugated goat anti-mouse/rabbit IgG.

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