Anti-STAT1 (phospho Y701) antibody [M135] (ab29045)

Protocol tips

Upstream tips	Protocol tips	Downstream tips
-These buffers may be stored at 4°C for several weeks or aliquoted and stored at -20°C for up to a year.	-Mouse (1:1000) -To reduce and denature your samples, boil each cell lysate in sample buffer at 100°C for 5 min. Lysates can be aliquoted and stored at -20°C for future use.

Upstream tips
-These buffers may be stored at 4°C for several weeks or aliquoted and stored at -20°C for up to a year.
Protocol tips
-Mouse (1:1000) -To reduce and denature your samples, boil each cell lysate in sample buffer at 100°C for 5 min. Lysates can be aliquoted and stored at -20°C for future use.

Publication protocol

Cells were collected and lysed on ice for 30 min in 1× radioimmunoprecipitation assay (RIPA) buffer plus 2× protease inhibitors. The lysate was centrifuged, and the supernatant was collected. Laemmli loading buffer with 0.05% BME was added to each sample. Samples were then loaded into the NuPAGE 10% (vol/vol) Bis-Tris SDS gels and 3-(N-morpholino)propanesulfonic acid buffer. The gels were run for 1 h at 180 V and were then transferred onto PVDF or nitrocellulose membranes using the Trans-Blot TURBO semidry transfer system from Bio-Rad.
The membrane was blocked for 1 h and then probed overnight with anti-GFP (ab6556; Abcam) at a dilution of 1:1,000, anti-IFNAR1 (MAR1-5A3) at a dilution of 1:500, STAT1 (ab92506; Abcam) at a dilution of 1:1,000, p-STAT1 (ab29045; Abcam) at a dilution of 1:1,000, or IFNAR1 (ab124764; Abcam) at a dilution of 1:1,000 in blocking solution at 4 °C overnight. The membrane was then washed three times for 10 min with Tris-buffered saline-tween (TBST). After the wash, the membrane was incubated with the corresponding Ab (HRP-conjugated) for 1 h at room temperature at a 1:5,000 dilution. The membrane was washed three times for 10 min in 1× TBST. Finally, the blots were overlaid with chemiluminescence ECL plus substrate and exposed to film.

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Manufacturer protocol

Download the product protocol from Abcam for Anti-STAT1 (phospho Y701) antibody [M135] (ab29045) below.

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