Cells were lysed in cold lysis buffer containing 5 mM NA3VO4, protease inhibitors (Complete, Roche, Bazel, Switzerland) and PhosSTOP (Roche). The proteins were separated on NuPAGE NOVEX Bis-Tris 4 to 12% gels (Invitrogen, Carlsbad, CA, USA). Western blot analysis was performed using antibodies against JAK3 (Cell Signaling, Danvers, MA, USA 3775), phospho-STAT5 (Y694) (Cell Signaling 9359); STAT5 (Invitrogen 335900), phospho-RanBP3 (Cell Signaling 9380); phospho-4E-BP1 (Cell Signaling 2855); pERK1/2 (Cell Signaling 9101), ERK1 (Santa Cruz, Dallas, TX, USA, Sc-93), pMEK1/2 (Cell Signaling 9154), MEK1/2 (Cell Signaling 9126) and β-actin (Sigma-Aldrich A1978); secondary antibodies were conjugated with horseradish peroxidase (GE Healthcare, Chicago, IL, USA). Bands were visualized using a cooled charge-coupled device camera (ImageQuant LAS-4000; GE Healthcare).
Stat5
Manufactured by Thermo Fisher Scientific
Sourced in United States
STAT5 is a laboratory equipment product designed for scientific research applications. It serves as a detection and quantification tool for the STAT5 protein, which is a key transcription factor involved in various cellular processes. The STAT5 product provides researchers with the necessary tools to analyze and study the expression and activity of the STAT5 protein in their research studies.
Automatically generated - may contain errors
Lab products found in correlation
Imagequant las 4000, by GE Healthcare (1 mentions)
P erk1 2, by Cell Signaling Technology (1 mentions)
Phosstop, by Roche (1 mentions)
β actin, by Merck Group (1 mentions)
Horseradish peroxidase, by GE Healthcare (1 mentions)
P mek1 2, by Cell Signaling Technology (1 mentions)
Phospho stat5 y694, by Cell Signaling Technology (1 mentions)
Phospho 4ebp1, by Cell Signaling Technology (1 mentions)
Mek1 2, by Cell Signaling Technology (1 mentions)
Las 4000, by Cytiva (1 mentions)
P stat3, by Thermo Fisher Scientific (1 mentions)
Polyvinylidene fluoride membrane, by Merck Group (1 mentions)
Enhanced chemiluminescence, by Thermo Fisher Scientific (1 mentions)
Gapdh, by Thermo Fisher Scientific (1 mentions)
Stat3, by Thermo Fisher Scientific (1 mentions)
Lysis buffer, by Cell Signaling Technology (1 mentions)
P jak1, by Thermo Fisher Scientific (1 mentions)
Pstat5, by Thermo Fisher Scientific (1 mentions)
2 protocols using stat5
1
Western Blot Analysis of Cell Signaling
2
Western Blot Analysis of JAK-STAT Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Total protein was obtained from cells or tissues using a lysis buffer (Cell Signaling Technology, California, USA), and protein levels were quantified using the BCA kit (Shanghai Ze Ye Biotechnology Co., Ltd, Shanghai, China). Subsequently, approximately 30 μg of protein was loaded and separated by 12% sodium dodecyl sulfate–polyacrylamide gel electrophoresis. The proteins were then transferred to a polyvinylidene fluoride membrane (Millipore, Massachusetts, USA), followed by incubation with 5% bovine serum albumin and incubation with the primary antibodies against JAK1 (1:800, Invitrogen, Massachusetts, USA), p-JAK1 (1:800, Invitrogen, Massachusetts, USA), JAK2 (1:800, Invitrogen, Massachusetts, USA), p-JAK1 (1:800, Invitrogen, Massachusetts, USA), STAT3 (1:800, Invitrogen, Massachusetts, USA), p-STAT3 (1:800, Invitrogen, Massachusetts, USA), STAT5 (1:800, Invitrogen, Massachusetts, USA), p-STAT5 (1:800, Invitrogen, Massachusetts, USA), and GAPDH (1:800, Invitrogen, Massachusetts, USA). Subsequently, the membranes were incubated with a secondary antibody (1:2000; Invitrogen, Massachusetts, USA). Finally, enhanced chemiluminescence (Invitrogen, Massachusetts, USA) was used for the visualization of bands, followed by quantification using the ImageJ software (National Institutes of Health, USA) [19 (link)].
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