STAT1 Monoclonal Antibody (15H3)

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	-Mouse (1:1000)

Protocol tips
-Mouse (1:1000)

Publication protocol

The harvested cells were lysed using lysis buffer (Beyotime Institute of Biotechnology, Haimen, China), and protein lysates (50 µg) were denatured in SDS sample buffer at 100°C for 10 min. The proteins were then separated by 10% SDS-PAGE and transferred onto nitrocellulose membranes. The membranes were blocked with 5% non-fat milk in TBST (25 mmol/l Tris-HCl, pH 7.5, 137 mmol/l NaCl, 2.7 mmol/l KCl and 0.05% Tween-20) for 1 h at 37°C, followed by incubation with mouse anti-signal transducer and activator of transcription (STAT1; 1:1,000; cat. no. MA1-037X; NeoMarkers; Thermo Fisher Scientific, Inc.), rabbit anti-Akt2 (1:1,000; cat. no. 3063; Cell Technology, Inc., Danvers, MA, USA) and mouse anti-β-actin (1:1,000; cat. no. sc-69879; Santa Cruz Biotechnology, Inc., Dallas, TX, USA) primary antibodies overnight at 4°C. The membrane was washed three times with TBST, followed by incubation with horseradish peroxidase-conjugated goat IgG secondary antibodies (1:1,000; cat. nos. sc-2004 and sc-2005; Santa Cruz Biotechnology, Inc.) for 2 h at room temperature. The antibodies were visualized using NBT/BCIP/buffer (1:1:50; Roche Diagnostics, Basel, Switzerland). The band intensities were quantified using ImageJ 1.48 software (National Institutes of Health, Bethesda, MD, USA).

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Manufacturer protocol

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