- 701110 - Invitrogen Antibodies STAT6 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of STAT6 in whole cell extracts of serum-starved HeLa cells treated with IFN-a (150 ng/mL, 15 min) using a STAT6 recombinant rabbit monoclonal antibody (Product # 701110) at a dilution of 2 µg/mL. Samples were detected using chemiluminescence (ECL). Results show a band at ~100 kDa.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent analysis of STAT6 in HeLa cells using a STAT6 recombinant rabbit monoclonal antibody (Product # 701110) followed by detection using an Alexa Fluor 488-conjugated goat anti-rabbit secondary antibody (green) (Image A). Nuclei were stained using DAPI (Image B) and actin stained with Alexa Fluor 594 phalloidin (red) (image C). Image D is a composite image showing cytoplasmic localization of STAT6.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemistry analysis of STAT6 showing staining in the cytoplasm and nucleus of paraffin-embedded human diffuse large B cell lymphoma (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 a STAT6 monoclonal antibody (Product # 701110) diluted in 3% BSA-PBS at a dilution of 1:100 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 prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of STAT6 showing staining in the cytoplasm and nucleus of paraffin-embedded human thymus 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 a STAT6 monoclonal antibody (Product # 701110) diluted in 3% BSA-PBS at a dilution of 1:100 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 prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of STAT6 showing staining in the cytoplasm and nucleus of paraffin-embedded mouse spleen 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 a STAT6 monoclonal antibody (Product # 701110) diluted in 3% BSA-PBS at a dilution of 1:100 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 prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Flow cytometry analysis of STAT6 was done on HeLa cells. Cells were fixed with 70% ethanol for 10 minutes, permeabilized with 0.25% Tritonª X-100 for 20 minutes, and blocked with 5% BSA for 1 hour at room temperature. Cells were labeled with ABfinityª STAT6 Recombinant Rabbit Monoclonal Antibody (701110, red histogram) or with rabbit isotype control (pink histogram) at 2 µg-4 µg/million cells in 2.5% BSA. After incubation at room temperature for 2-3 hours, the cells were labeled with Alexa Fluor¨ 488 Goat Anti-Rabbit Secondary Antibody (A11008) at a dilution of 1:400 for 30 minutes at room temperature. The representative 10,000 cells were acquired and analyzed for each sample using an Attune¨ Acoustic Focusing Cytometer. The purple histogram represents unstained control cells and the green histogram represents no-primary-antibody control.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: ChIP- qPCR analysis of STAT6 was performed with 3 µg/mL of the STAT6 Recombinant Rabbit Monoclonal Antibody (Product # 701110) on sheared chromatin from 2 million HeLa cells treated with IFN-gamma (50 ng/mL) for 1h using the MAGnify Chromatin Immunoprecipitation System (Product # 49-2024). 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 (Product # 4376600) with primers for the promoter of active AP-1 and COX2 gene, used as positive control targets, and the GAPDH 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.

701110