Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human placental tissue section (4µm) labelling CCR6 with ab93086. Tissue was treated with 15µg/ml Protenase for antigen retrieval and fixed with 4% PFA. A HRP-conjugated anti-mouse IgG was used as the secondary antibody (30 minutes at room temperature) followed by DAB color development.
IHC image of ab93086 staining in Human Tonsil 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 sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab93086, 1µg/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 Jurkat cells stained with ab93086 (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 (ab93086, 2µg/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 IgG2b [PLPV219] (ab91366, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in Jurkat cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.
Tang M et al. Expression and Prognostic Significance of Macrophage Inflammatory Protein-3 Alpha and Cystatin A in Nasopharyngeal Carcinoma. Biomed Res Int2015:617143 (2015).
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