Total protein from tissue samples or cells was extracted and protein concentrations were quantified using a BCA assay kit (Beyotime Biotechnology, China). Equal amounts of protein were separated using SDS-PAGE and electrophoretically transferred to PVDF membranes (Millipore). Antibodies against GAPDH (AF0006, Beyotime Biotechnology), β-actin (AF0003, Beyotime Biotechnology), and β-tubulin (10094-1-AP, Proteintech) were used as loading controls. Horseradish peroxidase (HRP)-conjugated secondary antibodies were used and the protein signals were visualized using ECL detection reagents (Millipore). Antibodies used were: USP11 (ab109232, Abcam), ERK1/2 (#4695, CST), phospho-ERK1/2 (#4370, CST), p38 (#8690, CST), p-p38 (#4511, CST), JNK (#9252, CST), p-JNK (#4668, CST), HA-tag (#3724, CST), Myc-tag (05-724,Sigma-Aldrich), Flag-tag (F3165, Millipore), PPP1CA (sc-271762, Santa Cruz Biotechnology), MEK1/2 (sc-81504, Santa Cruz Biotechnology) and p-MEK1/2 (sc-81,503, Santa Cruz Biotechnology).
Usp11
Manufactured by Abcam
USP11 is a deubiquitinating enzyme that plays a role in the regulation of protein stability. It is involved in the deubiquitination process, which removes ubiquitin molecules from target proteins, preventing their degradation by the proteasome.
Automatically generated - may contain errors
Lab products found in correlation
Mek1 2, by Santa Cruz Biotechnology (1 mentions)
P mek1 2, by Santa Cruz Biotechnology (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
Ecl detection reagent, by Merck Group (1 mentions)
Bca assay kit, by Beyotime (1 mentions)
Myc tag, by Merck Group (1 mentions)
β actin, by Beyotime (1 mentions)
Af0006, by Beyotime (1 mentions)
Af0003, by Beyotime (1 mentions)
H3k27me3, by Abcam (1 mentions)
Cyp24a1, by Merck Group (1 mentions)
Flag tag (flag), by Merck Group (1 mentions)
β actin, by Merck Group (1 mentions)
2 protocols using usp11
1
Quantitative Protein Analysis from Cells
2
Western Blot Analysis of Protein Expression
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Cells were lysed in RIPA buffer (50 mM Tris-HCl at pH 7.4, 150 mM NaCl, 1% NP-40, 1% sodium deoxycholate, 0.1% SDS) with cocktail (protease inhibitor). The protein supernatant was collected by centrifugation and the concentration was quantified. The proteins were transferred onto the PVDF (GE) membrane by electrophoresis and electrical transfer. Membranes were blocked with 5% non-fat milk and incubated with primary antibodies overnight at 4°C. On the second day, membranes were incubated with secondary antibodies for 1 h at RT. Images were obtained by X-ray film exposure. The antibodies used for western blot (WB) analysis were USP11 (Abcam, ab109232), LSH (Sigma, HPA063242), CYP24A1 (Sigma, HPA022261), GPX4 (Abcam, ab125066), SLC7A11 (CST, 12691), Multi-Ub (MBL, D058-3), Flag-tag (Sigma, F3165), HA-tag (CST, 3724), MYC-tag (MBL, M192-3S), His-tag (MBL, D291-3S), H3K27me3 (Abcam, ab192915), β-actin (Sigma, 4930), Tubulin (Sigma, T9026), H3 (CST, 60932), and horseradish peroxidase (HRP)-conjugated secondary antibodies (Jackson, 111-035-003; 115-035-003).
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