Recombinant Anti-JAK1 (phospho Y1022 + Y1023) antibody [EPR1899(2)] (ab138005)

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.	-Rabbit (1:500) -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
-Rabbit (1:500) -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

Each well of the plate was treated by 200 µl lysis buffer and maintained in an ice bath for 1 h. Sample solutions were then centrifuged for 15 min at 4°C. Protein concentration of the centrifuged supernatant was determined by Coomassie Brilliant Blue staining, and preserved at −80°C. Total protein (50 µg) from each sample was separated by electrophoresis. The samples were then transferred to PVDF membranes (90 V, 1.5 h at low temperature). Rabbit anti-human p-JAK (dilution: 1:500; cat. no.: ab138005), rabbit monoclonal p-STAT (dilution: 1:500; cat no.: ab32143) and mouse monoclonal PSMβ3 (dilution: 1:500; cat no.: ab128094), purchased from Abcam (Cambridge, MA, USA), were added after blocking and incubated at 4°C overnight, following four washes with phosphate-buffered saline (PBS). Horseradish peroxidase-conjugated goat anti-rabbit IgG (dilution: 1/2000; Abcam; cat no.: ab6721) was added to membranes and incubated at 4°C overnight. After three washes with PBS, the ECL enhanced chemiluminescence reagent kit was used for signal development. After thorough scanning of negative film, a semi-quantitative analysis was carried out on the bands by Shanghai Tianneng gel imaging processing system software. Data were normalized to levels of β-actin.

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Manufacturer protocol

Download the product protocol from Abcam for Recombinant Anti-JAK1 (phospho Y1022 + Y1023) antibody [EPR1899(2)] (ab138005) below.

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