Whole cell protein was extracted from frozen liver tissue, and total protein lysates were subjected to SDS-PAGE on 8–20% acrylamide gels, electro-transferred to polyvinylidene difluoride membranes (MilliporeSigma, Burlington, MA, United States), and probed overnight at 4°C in the presence of the following primary antibodies: anti-SIRT1 (1:1,000, 13161-1-ap, Proteintech Group, Inc., Rosemont, IL, United States), anti-PPARα (1:1,000, ab8934, Abcam, Cambridge, United Kingdom), and anti-SREBP (1:1,000, bs-1402R, Bioss Antibodies, Woburn, MA, United States). For protein detection, we used horseradish peroxidase-conjugated secondary antibodies (Wuhan ServiceBio Technology Co., Wuhan, China). Five samples per group per timepoint were assayed; protein levels were normalized to β-actin (1:3,000, GB12001, Wuhan ServiceBio Technology Co.). Densitometry analysis of protein bands was conducted using the open-source image processing software ImageJ (https://imagej.net/ImageJ ).
Bs 1402r
Manufactured by Bioss Antibodies
Sourced in United Kingdom
The Bs-1402R is a high-performance laboratory centrifuge designed for a variety of applications. It features a maximum speed of 14,000 rpm and a maximum RCF of 20,000 x g, making it suitable for a wide range of sample volumes and types. The unit is equipped with a brushless DC motor and electronic speed control for precise speed regulation. The Bs-1402R also includes a temperature control system to ensure samples are maintained at the desired temperature during the centrifugation process.
Automatically generated - may contain errors
Lab products found in correlation
Pvdf membrane, by Merck Group (2 mentions)
Ab8934, by Abcam (1 mentions)
Polyvinylidene difluoride membrane, by Merck Group (1 mentions)
Horseradish peroxidase conjugated secondary antibody, by Wuhan Servicebio Technology (1 mentions)
Gb12001, by Wuhan Servicebio Technology (1 mentions)
Casein blocker, by Bio-Rad (1 mentions)
Enhanced chemiluminescence, by Bio-Rad (1 mentions)
Bca kit, by Beyotime (1 mentions)
Protein lysis buffer, by Beyotime (1 mentions)
Stat3 4904p, by Cell Signaling Technology (1 mentions)
Ab8226, by Abcam (1 mentions)
Ab59493, by Abcam (1 mentions)
Bs 2442r, by Bioss Antibodies (1 mentions)
Bs 3550r, by Bioss Antibodies (1 mentions)
Bs 4509r, by Bioss Antibodies (1 mentions)
Bs 1630r, by Bioss Antibodies (1 mentions)
Bs 0295g, by Bioss Antibodies (1 mentions)
4 protocols using bs 1402r
1
Liver Protein Expression Analysis
2
Protein Extraction and Western Blot Analysis
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Mouse livers and hepatocytes were extracted with protein lysis buffer (Beyotime, P0013) supplemented with protease inhibitor cocktail. BCA Kit (Beyotime, P0009) was used to assess the protein concentration. Proteins (40 μg) were separated on 8–10% polyacrylamide precast SDS gels followed by blotting on PVDF membranes (Millipore Billerica, MA, USA). The membranes were blocked in 1×PBS 1% Casein Blocker (BioRad) diluted 1:10 for 1 h at room temperature and then incubated with antibodies against NONO (1:1000 dilution, Abcam, ab70335), TET2 (1:1000 dilution, Active motif, 61389), SREBP1(1:1000 dilution, Bioss, bs-1402R), CD36 (1:1000 dilution, Bioss, ET1701-24) and GAPDH (1:3000 dilution, Proteintech, 60004). HRP conjugate anti-Mouse IgG (1:10000 dilution, Proteintech, SA00001-1) and HRP conjugate anti-Rabbit IgG (1:10000 dilution, Proteintech, SA00001-2) were used as secondary antibody. Finally, the membranes were visualized with enhanced chemiluminescence (Bio-Rad, USA).
3
Western Blot Analysis of Transcription Factors
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Western blot experiments were performed as described before55 (link) using antibodies against SREBP1 (bs-1402R; BIOSS, Woburn, MA), LCN2 (AF1857; R&D Systems, Abingdon, UK), as well as phosho-c-JUN (3270), c-JUN (9165), p-STAT3 (Signal Transducers and Activators of Transcription 3) (9145), and STAT3 (4904P), all purchased from Cell Signaling Technology, Inc (Danvers, MA). Mouse anti–β-actin monoclonal antibodies (sc-47778; Santa Cruz Biotechnology, Inc, Dallas, TX) or Ponceau C–stained blots were used for loading controls. Semiquantitative analysis of obtained signals was performed using ImageJ software (National Institutes of Health, Bethesda, MD).56 (link)
4
Protein Expression Analysis in Transfected Cells
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Transfected cells were lysed in RIPA buffer containing 1% phenylmethylsulfonyl fluoride (PMSF). A BCA protein kit (Pierce, Thermo Fisher Scientific Inc., Waltham, MA, USA) was used to determine the total protein concentration. Protein samples were separated using 10% SDS-PAGE and transferred onto PVDF membranes (Millipore, Billerica, MA, USA). The membranes were incubated overnight at 4 °C with the following antibodies: anti-DGAT1 (ab189994, Abcam, Cambridge, UK), anti-DGAT2 (ab59493, Abcam, Cambridge, UK), anti-PPARγ (bs-4509R, Bioss, Beijing, China), anti-C/EBPα (bs-1630R, Bioss, Beijing, China), anti-SREBF1(bs-1402R, Bioss, Beijing, China), anti-Pax7 (ab61067, Abcam, Cambridge, UK), anti-MYOD (bs-2442R, Bioss, Beijing, China), and anti-MYOG (bs-3550R, Bioss, Beijing, China). Thereafter, the membranes were washed and incubated with horseradish peroxidase-conjugated goat anti-mouse or goat anti-rabbit IgG (bs-0295G; Bioss, Beijing, China) for 1 h. β-actin (ab8226, Abcam, Cambridge, UK) was used as an endogenous control. A grayscale intensity analysis was performed using ImageJ software (NIH).
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