ChIP- qPCR analysis of STAT2 was performed with 3 μg/mL of ab277764 on sheared chromatin from 2 million HeLa (Human epithelial cell line from cervix adenocarcinoma) cells treated with IFN-alpha (50 ng/mL) for 1h using the MAGnify Chromatin Immunoprecipitation System. Normal rabbit IgG (3 μg/mL) was used as a negative IP control. The purified DNA from each ChIP sample was analyzed by StepOnePlus Real-Time PCR System with primers for the promoter of active GLS1 gene, used as positive control target, and the OAS1 gene, used as negative control target. Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.

Immunohistochemistry analysis of STAT2 showing staining in the cytoplasm of paraffin-embedded human cervical carcinoma (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H₂O₂-methanol for 15 min at room temperature, washed with ddH₂O and PBS, and then probed with ab277764 diluted in 3% BSA-PBS at a dilution of 1/20 overnight at +4°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prepare for mounting.

All lanes : Anti-STAT2 antibody [1HCLC] (ab277764) at 3 µg/ml

Lane 1 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 2 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate
Lane 3 : A431 (Human epidermoid carcinoma cell line) whole cell lysate
Lane 4 : A549 (Human lung carcinoma cell line) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : HRP Goat anti-Rabbit IgG at 1/5000 dilution

Predicted band size: 97 kDa

Immunofluorescent analysis of STAT2 was done on 70% confluent log phase HeLa (Human epithelial cell line from cervix adenocarcinoma) cells. The cells were fixed with 4% paraformaldehyde for 15 minutes; permeabilized with 0.25% Triton X-100 for 10 minutes followed by blocking with 5% BSA for 1 hour at room temperature. The cells were incubated with ab277764 at 2 μg-4 μg in 1% BSA and incubated for 3 hours at room temperature and then labeled with Alexa Fluor 488 Goat anti-Rabbit IgG Secondary Antibody at a dilution of 1/400 for 30 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade Gold Antifade Mountant with DAPI. F-actin (Panel c: red) was stained with Alexa Fluor 594 Phalloidin. Panel d is a merged image showing translocated STAT2 in nucleus. Panel e shows cytoplasmic localization of STAT2. Panel f shows no primary antibody control. The images were captured at 20X magnification.

Immunohistochemistry analysis of STAT2 showing staining in the cytoplasm of paraffin-embedded human kidney tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H₂O₂-methanol for 15 min at room temperature, washed with ddH₂O and PBS, and then probed with ab277764 diluted in 3% BSA-PBS at a dilution of 1/20 overnight at +4°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prepare for mounting.

Anti-STAT2 antibody [1HCLC] (ab277764) at 2.5 µg/ml + MCF7 (Human breast adenocarcinoma cell line) whole cell lysate treated with IFN1a (150 ng/mL, 15 min)

Predicted band size: 97 kDa