LPS, GW4869 and PKH-67 were purchased from Sigma-Aldrich. The following antibodies were used: anti-CD68, anti-CD9, anti-CD63, anti-PTEN, anti-Alix, anti-Akt, anti-p-Akt, goat Anti-Rabbit IgG H&L (Alexa Fluor® 555) (Abcam), anti-p65, and anti-p-p65(Beyotime); miR-92a-3p mimic, mir-92a-3p inhibitor, control RNAs and the primers for miRNAs were all purchased from Guangzhou RiboBio.
Anti p p65
Manufactured by Beyotime
Sourced in United States
Anti-p-p65 is a primary antibody that specifically binds to the phosphorylated form of the p65 subunit of the NF-κB transcription factor. It can be used to detect and quantify the activated form of NF-κB in various experimental settings.
Automatically generated - may contain errors
Lab products found in correlation
Anti p65, by Beyotime (2 mentions)
Anti p65, by Proteintech (2 mentions)
Paeg gel fast preparation kit, by Ipsen (2 mentions)
Ecl femtolightsubstrate, by Ipsen (2 mentions)
Elisa ripa buffer, by Dingguo (2 mentions)
Anti β actin, by Proteintech (2 mentions)
Ripa buffer, by Dingguo (2 mentions)
Anti pten, by Abcam (1 mentions)
Anti alix, by Abcam (1 mentions)
Goat anti rabbit igg h l alexa fluor 555, by Abcam (1 mentions)
Mir 92a 3p inhibitor, by RiboBio (1 mentions)
Control rnas, by RiboBio (1 mentions)
Anti p akt, by Abcam (1 mentions)
Gw4869, by Merck Group (1 mentions)
Anti cd68, by Abcam (1 mentions)
Pkh67, by Merck Group (1 mentions)
Anti cd63, by Abcam (1 mentions)
Lipopolysaccharides (lps), by Merck Group (1 mentions)
Anti akt, by Abcam (1 mentions)
Anti cd9, by Abcam (1 mentions)
6 protocols using anti p p65
1
Exosome Regulation by miR-92a-3p
2
Protein Expression Analysis in RAW264.7 and BMSCs
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Western blot was used to detect the expression of different proteins in RAW264.7 and BMSCs. Anti-c-Fos (Beyotime, 1:2000), anti-NFATc1(ABclonal, 1:3000), anti-TRAP(Abcam, 1:5000), anti-cathepsin-K(Abcam, 1:2000), anti-TLR4(Sangon Biotech, 1:2000), anti-p-p65(Beyotime, 1:2000), anti-p65(ABclonal, 1:2000), anti-p-IκBα(ImmunoWay, 1:2000), anti-IκBα(ImmunoWay, 1:2000), anti-p-ERK (ABclonal, 1:2000), anti-ERK(ABclonal, 1:2000), anti-p-p38(ABclonal, 1:2000), anti-p38 (ABclonal, 1:2000), anti-p-JNK(ABclonal, 1:2000), anti-JNK(Abcam, 1:2000), anti-BMP-2(Abcam, 1:2000), anti-RUNX2(Abcam, 1:2000), anti-β-catenin(Abcam, 1:10,000), anti-p53(Abcam, 1:2000), anti-bax (Abcam, 1:10,000), anti-bcl-2(Abcam, 1:2000), anti-PPAR-γ(Solarbio, 1:2000), anti-β-actin(Abcam, 1:2000), goat anti-mouse IgG(Abcam, 1:5000), goat anti-rabbit IgG(Abcam, 1:5000) were used for protein analysis. ECL substrate kit (BL520B, biosharp) was used for visual analysis of proteins. The gray values of the bands were quantified by ImageJ software.
3
Immunoblotting and Flow Cytometry Protocols
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The following antibodies were used for immunoblotting: Cell Signaling Technology, anti-p38, anti-p-p38, anti-p65, anti-p-p65, anti-JNK, anti-p-JNK, anti-ERK, anti-p-ERK, anti-SHP-1, anti-p-SHP-1, anti-SHP-2, anti-p-SHP-2, anti-Lyn, anti-p-Lyn, the above antibodies were used at a 1:1,000 dilution; Beyotime Institute of Biotechnology, anti-β-actin (diluted at 1:1,000); HRP-labeled Goat anti-rabbit (1:3,000), HRP-labeled Rabbit anti-mouse (1:3,000). The following antibodies were used for flow cytometry: anti-human HLA-DR, anti-human CD14, anti-human CD11c, anti-human CD180, anti-human CD86, anti-mouse F4/80, anti-mouse CD11c, anti-mouse B220, anti-mouse CD40, anti-mouse CD86, anti-mouse CD180. All the antibodies used for flow cytometry were purchased from Biolegend and used at a 1:100 dilution. An isotype control was used for each antibody.
4
Western Blot Analysis of Inflammatory Markers
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The levels of the target proteins were determined by performing Western blotting as reported previously [49 (link),50 (link)]. Briefly, compound-treated cell lysates were heated with an SDS loading buffer for 10 min at 95 °C and separated by SDS-PAGE. The proteins were transferred and blotted on a polyvinylidene fluoride membrane (Millipore, Billerica, MA, USA) and incubated with primary antibodies obtained from Beyotime (anti-COX-2, anti-iNOS, anti-p-iκB, anti-iκB, anti-p65, anti-p50, or anti-GAPDH) or obtained from Cell Signaling Technology (anti-p-p65, Beverly, MA, USA) and an HRP-conjugated secondary antibody purchased from Beyotime. The proteins were visualized by chemiluminescence according to the Beyotime ECL kit’s protocol and analyzed using ImageJ 1.53k software.
5
Western Blot and ELISA for Inflammatory Markers
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Samples were lysed using RIPA buffer (Dingguo, China) based on provided directions.The total protein was separated by PAEG gel fast preparation kit (Epizyme, China) and transferred to polyvinylidene uoride (PVDF) membranes. The membrances were blocked with bovine serum albumin for 2 h at room temperature. Then, primary antibodies against rabbit p65 (anti-P65, Proteintech, USA), Phospho-NF-κB p65 (Ser536) (anti-p-p65, Beyotime), and β-actin (anti-β-actin, Proteintech) were incubated overnight at 4°C. Followed, washed it with TBST and incubated with goat anti-rabbit secondary for 1 h. Finally, the membranes were exposed using ECL FemtoLightSubstrate (Epizyme). The immunoreactive protein bands were quantifed using ImageJ software.
Enzyme-linked immunosorbent assay (ELISA)
RIPA buffer (Dingguo) was used to prepare proteins from cell lysates and brain homogenates. Levels of IL-1β, IL-6, TNF-α, and iNOS in these samples were then measured using commercial ELISA kits (Beyotime) based on provided directions.
Enzyme-linked immunosorbent assay (ELISA)
RIPA buffer (Dingguo) was used to prepare proteins from cell lysates and brain homogenates. Levels of IL-1β, IL-6, TNF-α, and iNOS in these samples were then measured using commercial ELISA kits (Beyotime) based on provided directions.
6
Western Blot and ELISA for Inflammatory Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Samples were lysed using RIPA buffer (Dingguo, China) based on provided directions.The total protein was separated by PAEG gel fast preparation kit (Epizyme, China) and transferred to polyvinylidene uoride (PVDF) membranes. The membrances were blocked with bovine serum albumin for 2 h at room temperature. Then, primary antibodies against rabbit p65 (anti-P65, Proteintech, USA), Phospho-NF-κB p65 (Ser536) (anti-p-p65, Beyotime), and β-actin (anti-β-actin, Proteintech) were incubated overnight at 4°C. Followed, washed it with TBST and incubated with goat anti-rabbit secondary for 1 h. Finally, the membranes were exposed using ECL FemtoLightSubstrate (Epizyme). The immunoreactive protein bands were quantifed using ImageJ software.
Enzyme-linked immunosorbent assay (ELISA)
RIPA buffer (Dingguo) was used to prepare proteins from cell lysates and brain homogenates. Levels of IL-1β, IL-6, TNF-α, and iNOS in these samples were then measured using commercial ELISA kits (Beyotime) based on provided directions.
Enzyme-linked immunosorbent assay (ELISA)
RIPA buffer (Dingguo) was used to prepare proteins from cell lysates and brain homogenates. Levels of IL-1β, IL-6, TNF-α, and iNOS in these samples were then measured using commercial ELISA kits (Beyotime) based on provided directions.
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