IHC image of FOXP3 staining in a section of formalin-fixed paraffin-embedded normal human spleen* performed on a Leica BOND^TM 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 20mins. The section was then incubated with ab191416, 5ug/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. The inset secondary-only control image is taken from an identical assay without primary antibody.

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.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

All lanes : Anti-FOXP3 antibody [EPR15038-69] (ab191416) at 5 µg/ml

Lane 1 : HEK293T cell lysate
Lane 2 : HEK293T cell lysate overexpressing Human FOXP3

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 47 kDa

This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using Licor blocking buffer before being incubated with ab10563. Antibody binding was detected using Goat anti Rabbit IR680 at a 1:10,000 dilution for 1hr at room temperature and then imaged using the Licor Odyssey CLx.

IHC image of FOXP3 staining in a section of formalin-fixed frozen normal human tonsil* performed on a Leica BOND^TM system using the standard protocol F. The section was then incubated with ab191416, 5ug/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. The inset secondary-only control image is taken from an identical assay without primary antibody.

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.

 

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

IHC image of FOXP3 staining in a section of formalin-fixed paraffin-embedded normal human tonsil* performed on a Leica BOND^TM 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 20mins. The section was then incubated with ab191416, 5ug/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. The inset secondary-only control image is taken from an identical assay without primary antibody.

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.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

Immunohistochemical analysis of paraffin-embedded Human Hodgkin lymphoma tissue labeling FOXP3 with ab191416 at 1/250 dilution (0.9 μg/ml) followed by pre-diluted HRP Polymer for Rabbit IgG secondary antibody and counter-stained with Hematoxylin.
Inset: Negative control: using PBS instead of primary antibody.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.