Western blot - AP2 alpha antibody (ab52222)

All lanes : Anti-AP2 alpha antibody - ChIP Grade (ab52222) at 1/500 dilution

Lane 1 : extracts from COLO205 cells,
Lane 2 : extracts from COLO205 cells, with immunizing peptide


Predicted band size : 48 kDa
Observed band size : 48 kDa

Electrophoretic Mobility Shift Assay - AP2 alpha antibody (ab52222)

Antibody anti-AP-2 alpha (ab52222) was used in an Electrophoretic Mobility Shift Assay (EMSA) to supershift the protein-DNA complex.

Radiolabelled, double-stranded DNA oligonucleotides (10.000 cpm per lane) harbouring a binding site for AP-2 alpha were incubated with each 2 µg of nuclear extract (NE) from HeLa and Caski cells, respectively. Samples were incubated for 30 minutes at room temperature to allow the formation of protein-DNA complexes. 2 µg of anti-AP-2 alpha antibody were added to the samples (as indicated) and incubated for further 60 minutes at 4ºC. Samples were separated in a 5.5% PAGE for 30 minutes at 280 V and further 75 minutes at 350 V. The Gel was dried under vacuum and for autoradiography a X-ray film was exposed with an intensifying screen for 2 days at -80ºC. Specific protein-DNA complexes were quantitatively supershifted with antibody anti-AP-2 alpha (ab52222), verifying the binding of AP-2 alpha to the DNA oligonucleotide. Rating: ***** This image was kindly provided by Dr Lars Krueger.

Electrophoretic Mobility Shift Assay - AP2 alpha antibody (ab52222)

Antibody anti-AP-2 alpha (ab52222) was used in an Electrophoretic Mobility Shift Assay (EMSA) to supershift the protein-DNA complex.

Radiolabelled, double-stranded DNA oligonucleotides (10.000 cpm per lane) harbouring a binding site for AP-2 alpha were incubated with each 2 µg of nuclear extract (NE) from HeLa and Caski cells, respectively. Samples were incubated for 30 minutes at room temperature to allow the formation of protein-DNA complexes. 2 µg of anti-AP-2 alpha antibody were added to the samples (as indicated) and incubated for further 60 minutes at 4ºC. Samples were separated in a 5.5% PAGE for 30 minutes at 280 V and further 75 minutes at 350 V. The Gel was dried under vacuum and for autoradiography a X-ray film was exposed with an intensifying screen for 2 days at -80ºC. Specific protein-DNA complexes were quantitatively supershifted with antibody anti-AP-2 alpha (ab52222), verifying the binding of AP-2 alpha to the DNA oligonucleotide. Rating: ***** This image was kindly provided by one of our customers.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-AP2 alpha antibody - ChIP Grade (ab52222)

IHC image of AP2 alpha staining in human breast carcinoma formalin fixed paraffin embedded tissue section, 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 20 mins. The section was then incubated with ab52222, 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.