重组Anti-Estrogen Receptor alpha抗体[EPR4097] (ab108398)

ChIC/CUT&RUN was performed using a pAG-MNase at a final concentration of 700 ng/mL, 2.5 x 10^5 MCF7 (Human breast adenocarcinoma epithelial cell) cells treated with phenol red free medium and 5% charcoal stripped FBS for 3 days than treated with β-estradiol (10 nM 45 min) and 5 µg of ab108398 [EPR4097]. The resulting DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 10 million reads. The negative IgG control ab172730 is also shown. The University of Geneva owns patents relevant to ChIC (Chromatin Immuno-Cleavage) methods.

ChIC/CUT&RUN was performed using a pAG-MNase at a final concentration of 700 ng/mL, 2.5 x 10^5 MCF7 (Human breast adenocarcinoma epithelial cell) cells treated with phenol red free medium and 5% charcoal stripped FBS for 3 days than treated with β-estradiol (10 nM 45 min) and 5 µg of ab108398 [EPR4097]. The resulting DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 10 million reads. The negative IgG control ab172730 is also shown. The University of Geneva owns patents relevant to ChIC (Chromatin Immuno-Cleavage) methods.

ChIC/CUT&RUN was performed using a pAG-MNase at a final concentration of 700 ng/mL, 2.5 x 10^5 MCF7 (Human breast adenocarcinoma epithelial cell) cells treated with phenol red free medium and 5% charcoal stripped FBS for 3 days than treated with β-estradiol (10 nM 45 min) and 5 µg of ab108398 [EPR4097]. The resulting DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 10 million reads. The negative IgG control ab172730 is also shown. The University of Geneva owns patents relevant to ChIC (Chromatin Immuno-Cleavage) methods.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast tissue labelling Estrogen Receptor alpha with purified ab108398 at 1/250. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. A prediluted HRP-polymer conjugated anti-rabbit IgG was used as the secondary antibody. Counterstained with Hematoxylin.

Blocking and diluting buffer: 5% NFDM/TBST

Blocking/Diluting buffer and concentration: 5% NFDM/TBST

Lane 1: Recombinant Human Estrogen Receptor beta (aa 1 to 323) protein, (Cat#: ab158385)
There is no cross-reactivity between ab108398 and ER beta protein.

Blocking buffer and concentration: 5% NFDM/TBST.

Diluting buffer and concentration: 5% NFDM /TBST.

Immunocytochemsitry/Immunofluorescence analysis of MCF-7 cells labelling Estrogen Receptor alpha (green) with purified ab108398 at 1/200. Cells were fixed with 4% paraformaldehyde. An Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/200) was used as the secondary antibody. Counterstained with DAPI (blue).

ab108398 staining Estrogen Receptor alphain the human cell line MCF-7 (human breast carcinoma) by intracellular flow cytometry. Cells were fixed with 4% paraformaldehyde, permeabilized with 90% methanol and the sample was incubated with the primary antibody at a dilution of 1/20. A goat anti rabbit IgG (Alexa Fluorr^® 488) at a dilution of 1/2000 was used as the secondary antibody. 

Isoytype control: Rabbit monoclonal IgG (Black) 

Unlabelled control: Cell without incubation with primary antibody and secondary antibody (Blue) 

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast ductal infiltrating carcinoma tissue labelling Estrogen Receptor alpha with unpurified ab108398.

Immunohistochemistry (Formalin/PFA-fixed parffin-embedded sections) analysis of human normal tonsil tissue. Unpurified ab108398 shows negative staining.

Immunohistochemistry (Formalin/PFA-fixed parffin-embedded sections) analysis of human colonic adenocarcinoma tissue. Unpurified ab108398 shows negative staining.

Immunohistochemistry (Formalin/PFA-fixed parffin-embedded sections) analysis of human lung adenocarcinoma tissue. Unpurified ab108398 shows negative staining.

Immunohistochemistry (Formalin/PFA-fixed parffin-embedded sections) analysis of human cervical carcinoma tissue. Unpurified ab108398 shows negative staining.

IHC image of Estrogen Receptor alpha staining in a section of frozen human uterus* performed on a Leica Biosystems BOND^® RX instrumen using the standard protocol. The section was fixed in 10% paraformaldehyde (10 min) prior to staining. The section was incubated with ab108398, 5 ug/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 Estrogen Receptor alpha staining in a section of frozen human cervix* performed on a Leica Biosystems BOND^® RX instrumen using the standard protocol. The section was fixed in 10% paraformaldehyde (10 min) prior to staining. The section was incubated with ab108398, 5 ug/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.

Negative control image: IHC image of Estrogen Receptor alpha staining in a section of frozen human hippocampus* performed on a Leica Biosystems BOND^® RX instrumen using the standard protocol. The section was fixed in 10% paraformaldehyde (10 min) prior to staining. The section was incubated with ab108398, 5 ug/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.