Primary antibodies were obtained as follows: LC3B, tubulin, and MuRF1 antibodies were acquired from Novus Biologicals (Centennial, CO, USA), Abcam (Cambridge, UK), and Santa Cruz Biotechnology (Dallas, TX, USA), respectively; mTOR, pS2448-mTOR, Akt, pS473-Akt, ribosomal S6, pS235/236-ribosomal S6, 4EBP1, pS65-4EBP1, FoxO1, pS256-FoxO1, Atrogin-1, ULK1, pS757-ULK1, AMPK, and pT172-AMPK were obtained from Cell Signaling Technology (Danvers, MA, USA). All secondary antibodies were procured from Jackson Immuno Research Laboratories, Inc. (West Grove, PA, USA). The intercostal, diaphragm, and gastrocnemius muscles from male Sprague Dawley rats, either 6-month-old or 20-month-old were obtained from the Aging Tissue Bank (Pusan University, Pusan, Korea) [49 (link)].
Murf1
Manufactured by Novus Biologicals
Sourced in United States, United Kingdom
MuRF1 is a ubiquitin ligase that plays a role in the regulation of muscle protein turnover. It is involved in the ubiquitination and subsequent degradation of muscle proteins by the proteasome.
Automatically generated - may contain errors
Lab products found in correlation
Mafbx, by Santa Cruz Biotechnology (2 mentions)
Tubulin, by Novus Biologicals (1 mentions)
Ps473 akt, by Cell Signaling Technology (1 mentions)
Atrogin 1, by Cell Signaling Technology (1 mentions)
Ps2448 mtor, by Cell Signaling Technology (1 mentions)
Ps256 foxo1, by Cell Signaling Technology (1 mentions)
Pt172 ampk, by Cell Signaling Technology (1 mentions)
4e bp1, by Cell Signaling Technology (1 mentions)
Foxo1, by Cell Signaling Technology (1 mentions)
Murf1, by Santa Cruz Biotechnology (1 mentions)
Total yap, by Santa Cruz Biotechnology (1 mentions)
Myostatin, by Abcam (1 mentions)
Tumor necrosis factor (tnf), by Santa Cruz Biotechnology (1 mentions)
Immobilon, by Merck Group (1 mentions)
Foxo1, by Abcam (1 mentions)
Calpain, by Santa Cruz Biotechnology (1 mentions)
P iκbα, by Santa Cruz Biotechnology (1 mentions)
Protease inhibitor cocktail, by Merck Group (1 mentions)
3 protocols using murf1
1
Molecular Markers of Muscle Aging
2
Comprehensive Immunoblotting Assay Protocols
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For immunoblotting, primary antibodies against S6K1 (Thr389, #9234; total #2708), mTOR (Ser2448, #2971; total, #2983), 4E‐BP1 (Thr37/46, #2855; total, #9644), eEF2 (Thr56, #2331; total, #2332), JNK (Thr183/Tyr185, #4668), SMAD2 (Ser245/250/255, #3104; total, #5339), TAZ (Ser89, #59971), ACC (Ser79, #3661; total, #3676), AMPK (Thr172, #4188; total, #2532), ULK1 (Ser317, #12753; total, #8054), LC3b (#2775), GABARAP (#13733), BNIP3 (#44060) TSC2 (Ser1387, #5584; total, #3635) PRAS40 (Thr246, #13175; total, #2691) and p38 (Thr180/Thr182, #4511; total, #8690), were purchased from CST (Beverly, MA, USA). Primary antibody for total YAP (sc‐101199), total TAZ (sc‐293183), total JNK (sc‐7345), MuRF‐1 (#sc‐398608), MAFbx (#sc‐16806) and UBR5 (#sc‐515494) were purchased from Santa Cruz Biotechnology (Heidelberg, Germany). Primary antibody for YAP (Ser127, #PA5‐17481) was purchased from Thermo Scientific. Primary antibody for REDD1 (#63059) was purchased from Abcam (Cambridge, UK). Primary antibody for HIF‐1α (#NB100‐134) was purchased from Novus Biologicals (Abingdon, UK).
All primary antibodies were diluted 1:1000 except for the total eEF2 and total 4E‐BP1, which were diluted 1:2000, and total JNK, total YAP, total TAZ, UBR5, MuRF‐1 and MAFbx which were diluted 1:500. Secondary anti‐rabbit (#7074; 1:10 000) and secondary anti‐mouse (#7076; 1:10 000) were purchased from CST.
All primary antibodies were diluted 1:1000 except for the total eEF2 and total 4E‐BP1, which were diluted 1:2000, and total JNK, total YAP, total TAZ, UBR5, MuRF‐1 and MAFbx which were diluted 1:500. Secondary anti‐rabbit (#7074; 1:10 000) and secondary anti‐mouse (#7076; 1:10 000) were purchased from CST.
3
Protein Extraction and Analysis from Tumors
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Proteins from tumors were extracted with a buffer solution (20 mM Tris-HCl, pH 7.5, 2 mM ATP, 5 mM MgCl2, 1 mM dithiothreitol (DTT), and 5 μL of a protease inhibitor cocktail (Sigma)). Proteins (20 μg/lane) were separated on a 12.5% polyacrylamide gel (a precast SDS gel (Bio-Rad)) and then transferred to a polyvinylidene difluoride membrane (Immobilon, Millipore). Proteins were determined using antibodies against mouse IL-6 (1:200, Abcam), TNF (1:200, Santa Cruz Biotechnology), Myostatin (1:100, Abcam), MAFbX (1:200, Santa Cruz Biotechnology), MuRF 1 (1:100, Novus), Cathepsina L (1:100, Abcam), Calpain (1:100, Santa Cruz Biotechnology), p-IκB-α (1:100, Santa Cruz Biotechnology) and FOXO1 (1:200, Abcam). The antibodies then were stripped off the membrane and re-probed with a specific antibody against β-actin (1:5000, Novus Biologicals). The intensity was quantified using the Fotodyne Image analysis System (Fotodyne, Hartland, WI, USA) and the TotalLab software (Nonlinear Dynamics, Durham, NC, USA).
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