Subsequent to the indicated treatments, protein of cells was harvested with RIPA buffer (Beyotime, Shanghai, China) supplemented with PMSF (Beyotime) and determined using a bicinchoninic acid assay kit (Beyotime). Proteins were separated with 10% or 12% SDS-PAGE gels and transferred PVDF membranes (Merck Millipore, Billerica, MA, USA), which were blocked with 5% non-fat milk for 1 h at room temperature. The membranes were incubated with primary antibody at 4°C overnight and then blotted with secondary antibody for 2 h at room temperature. The bands were detected using enhanced chemiluminescence (Merck Millipore). The primary antibodies against p62 (1:1000), LC3 (1:1000), and β-actin (1:1000) were obtained from Cell Signaling Technology (Danvers, MA, USA).
Primary antibodies against p62
Manufactured by Cell Signaling Technology
Sourced in United States
Primary antibodies against p62. p62 is a multifunctional protein that plays a role in various cellular processes. These antibodies can be used to detect and analyze p62 in biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Enhanced chemiluminescence, by Merck Group (1 mentions)
β actin, by Cell Signaling Technology (1 mentions)
Ripa buffer, by Beyotime (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
Phenyl methyl sulfonyl fluoride (pmsf), by Beyotime (1 mentions)
Bicinchoninic acid assay kit, by Beyotime (1 mentions)
Polyvinylidene fluoride (pvdf), by Merck Group (1 mentions)
Enhanced chemiluminescence detection system, by Thermo Fisher Scientific (1 mentions)
Gapdh, by Thermo Fisher Scientific (1 mentions)
Protease inhibitor cocktail, by Roche (1 mentions)
Gapdh, by Santa Cruz Biotechnology (1 mentions)
Ripa buffer, by Wanlei (1 mentions)
P atk, by Cell Signaling Technology (1 mentions)
Glutaraldehyde, by Yuanye Bio-Technology (1 mentions)
Il 1β, by Proteintech (1 mentions)
Primary antibodies against inos, by Proteintech (1 mentions)
3 methyladenine 3 ma, by Merck Group (1 mentions)
P pi3k, by Cell Signaling Technology (1 mentions)
Rutin, by Yuanye Bio-Technology (1 mentions)
3 protocols using primary antibodies against p62
1
Western Blot Analysis of Autophagy Markers
2
Western Blot Analysis of Autophagy Markers
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Total protein was extracted from treated cells with RIPA buffer (Wanlei Biotechnology, Shanghai, China) containing a protease inhibitor cocktail (Roche, Mannheim, Germany). The protein samples (30 μg per lane) were loaded onto 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels and electro-transferred onto polyvinylidene difluoride membranes (PVDF) (Millipore, Billerica, MA, USA). Then, the membranes were saturated with 5% non-fat milk in Tris-buffered saline with Tween (TBST) for 2 h at room temperature and separately incubated at 4 °C overnight with primary antibodies against p62 (1 : 1000 dilution; Cell Signaling Technology, Danvers, MA, USA), autophagy related protein-5 (Atg-5) (1 : 1000 dilution; Cell Signaling Technology), LC3 (1 : 1000 dilution; Abcam, Cambridge, MA, USA), TRIM16 (1 : 500 dilution; Abcam) and GAPDH (1 : 2000; Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA), followed by horseradish peroxidase (HRP)-conjugated secondary antibody IgG (1 : 2000 dilution; Cell Signaling Technology). The protein signals were visualized using an enhanced chemiluminescence detection system (Thermo Fisher Scientific, Inc.), with GAPDH as a loading control.
3
Rutin Attenuates Ox-LDL-Induced Macrophage Activation
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RAW264.7 mouse macrophages were obtained from the cell bank of the Chinese Academy of Sciences. Rutin was obtained from Shanghai Yuanye Biotechnology Co., Ltd., B20771, Shanghai, China, while ox-LDL was purchased from Guangzhou Yiyuan Biotechnology Co., Ltd., YB-002, Guangzhou, China. Glutaraldehyde at a concentration of 2.5% (Shanghai Yuanye Biotechnology Co., Ltd., R20510, Shanghai, China) was prepared. The primary antibodies against INOS, Arg1, IL-1β, and Mcp1 were obtained from Proteintech (Wuhan, China), while primary antibodies against P62, LC3, PI3K, p-PI3K, ATK, and p-ATK were bought from Cell Signaling Technology (Boston, MA, USA). Sigma-Aldrich provided the 3-methyladenine (3-MA) (St. Louis, MO, USA).
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