Lipopolysaccharides (LPS) (Escherichia coli O111:B4) were purchased from Sigma Aldrich. ON-TARGETplus SMARTpool siRNA and single siRNA targeting mouse Ruvbl2 and Ruvbl1 were obtained from Dharmacon. For immunoblotting and ChIP assay, the following antibodies were used: mouse monoclonal anti-Reptin 52 (RUVBL2) (Santa Cruz), mouse anti-TIP49A (RUVBL1) (Abcam), mouse anti-β-Actin (Sigma-Aldrich), rabbit anti-p38/MAPK (Cell Signaling Technology), rabbit anti-p-p38/MAPK (T180/Y182; Cell Signaling Technology), rabbit anti-p44/42 (ERK1/2; Cell Signaling Technology), rabbit anti-p-p44/42 MAPK (ERK1/2; Thr202/Tyr204 (Cell Signaling Technology), rabbit anti-SAPK/JNK (Cell Signaling Technology), rabbit anti-p-SAPK/JNK (Thr183/Tyr185) (Cell Signaling Technology), rabbit anti-IκBα (Cell Signaling Technology), rabbit anti-Stat1 (Cell Signaling Technology), rabbit anti-p-Stat1(S727) (Cell Signaling Technology), rabbit anti-p-Akt (S473) (Cell Signaling Technology), mouse anti-LAMIN B1 (Santa Cruz), mouse anti-α-tubulin (Sigma-Aldrich), rabbit anit-p50 (Abcam), rabbit anti-H3K4Me3 (Cell Signaling Technology), rabbit anti-H4K20me3 (Santa Cruz Biotechnology), and rabbit anti-p50 (Cell Signaling Technology) antibodies. CB-6644 was obtained from MedChemExpress.
Rabbit anti iκbα
Manufactured by Cell Signaling Technology
Sourced in United States
Rabbit anti-IκBα is a primary antibody that specifically recognizes the IκBα protein. IκBα is an inhibitor of the NF-κB transcription factor, which plays a crucial role in cellular signaling pathways. This antibody can be used to detect and quantify IκBα levels in various experimental systems.
Automatically generated - may contain errors
Lab products found in correlation
Rabbit anti nf κb p65, by Cell Signaling Technology (4 mentions)
Rabbit anti p iκbα, by Cell Signaling Technology (4 mentions)
Mouse anti β actin, by Merck Group (3 mentions)
Mouse anti α tubulin, by Merck Group (3 mentions)
Rabbit anti phospho nf κb p65 ser536, by Cell Signaling Technology (3 mentions)
Rabbit anti p p65, by Cell Signaling Technology (3 mentions)
Rabbit anti p38, by Cell Signaling Technology (3 mentions)
Rabbit anti p p38, by Cell Signaling Technology (3 mentions)
Rabbit anti p38 mapk, by Cell Signaling Technology (2 mentions)
Rabbit anti β actin, by Cell Signaling Technology (2 mentions)
Rabbit anti tak1, by Cell Signaling Technology (2 mentions)
Rabbit anti p65, by Cell Signaling Technology (2 mentions)
Mouse anti phospho iκbα, by Cell Signaling Technology (2 mentions)
Rabbit anti phospho iκbα, by Cell Signaling Technology (2 mentions)
Mouse anti caspase 8, by Enzo Life Sciences (2 mentions)
Rabbit anti nik, by Cell Signaling Technology (2 mentions)
Rabbit anti erk1 2, by Cell Signaling Technology (2 mentions)
Rabbit anti p erk1 2, by Cell Signaling Technology (2 mentions)
Mouse anti gapdh, by Santa Cruz Biotechnology (2 mentions)
Odyssey imaging system, by LI COR (2 mentions)
26 protocols using rabbit anti iκbα
1
LPS-Induced RUVBL1/2 Signaling Pathway
2
Western Blot Analysis of Signaling Proteins
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After 48 h transfection, cultured SGC-7901 cells were washed with cold PBS once and lysed with lysis buffer (0.5% NP40). Equal quantities of protein (30 µg) were separated by 10% SDS-PAGE and transferred to nitrocellulose membranes. Membranes were blocked with 5% skimmed milk and incubated with the following primary antibodies, diluted 1:1,000, overnight at 4°C: Rabbit anti-TAK1 (catalog no. 4505), rabbit anti-IκBα (catalog no. 4812), anti-rabbit B-cell lymphoma 2 (Bcl-2; catalog no. 4223), rabbit anti-β-actin (catalog no. 4970) and rabbit anti-Flag (catalog no. 14793), all purchased from Cell Signaling Technology, Inc. Blots were washed with TBS containing Tween 20 (TBST) three times for 10 min each time. Subsequently, membranes were incubated with a horseradish peroxidase-conjugated goat anti-rabbit antibody (1:3,000; catalog no. A00098; GenScript, Piscataway, NJ, USA) for 2 h at room temperature and washed with TBST three times for 10 min each time. Protein bands were visualized using Enhanced Chemiluminescence Plus (Thermo Fisher Scientific, Inc.) and scanned by Typhoon™ FLA 9500 (GE Healthcare Life Sciences, Chalfont, UK). Densitometry was quantified using Image J software (National Institutes of Health, Bethesda, MD, USA) and normalized to β-actin.
3
Ang-(1-7) Modulates Inflammation and Oxidative Stress
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LPS was purchased from Sigma-Aldrich (St. Louis, MO, United Ststes). Ang-(1-7) was purchased from Gill Biochemistry (Shanghai, China). Ang II enzyme-linked immunosorbent assay (ELISA) kit, Ang II polyclonal antibody, TNFα, IL-1β, and IL-6 were purchased from Ray Biotech (Guangzhou, China). rabbit anti-Phospho-NF-κB-p65 (Ser536) and rabbit anti-IκBα were purchased from Cell Signaling Technology (Boston, United Ststes). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and bovine serum albumin (BSA) were obtained from Jiangsu Kaiji Biotechnology (Nanjing, China). Malondialdehyde (mda), superoxide dismutase (sod), and total antioxidant capacity (TAOC) detection kits were provided by Nanjing Institute of Bioengineering (Nanjing, China).
4
Western Blot Analysis of Liver Proteins
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Total protein of 100 mg snap-frozen liver tissues was extracted by RIPA lysis buffer and protein concentration was determined by BCA assay kit (Solarbio, Beijing, China) based on the manufacturer's protocols. 40 μg protein was loaded in 10% SDS-PAGE gel and transferred to PVDF membranes. Rabbit anti-IκBα (#4812, 1 : 1000, Cell Signaling Technology, Inc., Danvers, MA, USA), anti-NF-κB P65 (#8242, 1 : 1000, Cell Signaling Technology, Inc., Danvers, MA, USA), anti-CYP2E1 (ab28146, 1 : 2000, Abcam, Cambridge, UK), anti-Histone H3 (#4499, 1 : 2000, Cell Signaling Technology, Inc., Danvers, MA, USA), and anti-GAPDH (ab9484, 1 : 2500, Abcam, Cambridge, UK) antibodies were used and goat anti-rabbit IgG H&L (HRP) (ab205718, 1 : 5000, Abcam, Cambridge, UK) as the secondary antibody. The membranes were visualized by ECL reagents, and the protein bands were detected by G:BOX gel imaging system (Syngene, Cambridge, UK).
5
Western Blot Analysis of Signaling Pathways
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The cells were washed twice with ice‐cold PBS and solubilized in lysis buffer supplemented with phosphatase (Roche) and protease inhibitor cocktails (Cell Signaling Technology). Cells were further lysed by sonication (TOMY Seiko), and lysates were clarified by centrifugation at 14 000 RCF for 10 min at 4°C. Supernatants were collected, and protein concentrations were determined with the Pierce BCA Protein Assay Kit (Thermo Fisher Scientific). The supernatants were boiled in samples buffer containing 0.125 M Tris–HCl (pH 6.8), 40% glycerol, 4% sodium dodecyl sulfate (SDS), 0.2 M dithiothreitol, and 0.01% bromophenol blue, subjected to SDS‐polyacrylamide gel electrophoresis and transferred to a nitrocellulose membrane. Membranes were probed with primary antibody followed by the appropriate horseradish peroxidase‐conjugated secondary antibody. Using Immobilon Western Chemiluminescent HRP Substrate (Millipore), bands were detected by chemiluminescence and imaged using a LAS‐4000 Imaging System (Fujifilm). The antibodies used were as follows: Rabbit anti‐Phospho‐NF‐κB p65 (Cell Signaling; #3033), Rabbit anti‐NF‐κB p65 (Cell Signaling; #8242), Rabbit anti‐IκBα (Cell Signaling; #4812), Rabbit anti‐Phospho‐Smad1/5 (Cell signaling; #9516), Rabbit anti‐Smad1 (Cell signaling; #6944), Anti‐GAPDH‐HRP (MBL; #M171‐7), Goat Anti‐Rabbit HRP (Jackson ImmunoResearch; #111‐035‐144).
6
Western Blot Analysis of Signaling Proteins
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The proteins were separated by SDS-PAGE, which was preformed according to the instructions, and the separated proteins were transferred to a PVDF membrane. The membrane was blocked with skim milk. After washing with TBS-T, the membrane was incubated with the primary antibody at 4°C overnight. After thorough washing with TBS-T, the secondary antibody was applied to the membrane and detected using a chemiluminescence detection kit and related instruments. The primary antibodies used were mouse anti-CGRP (1 : 500, Santa Cruz Biotechnology, CA, USA), mouse anti-SP (1 : 500, Santa Cruz Biotechnology, CA, USA), anti-Oprk1 (1 : 500, Sigma-Aldrich, St. Louis, MO, USA), rabbit anti-p-P65 (1 : 1000, Cell Signaling Technology, Danvers, MA, USA), rabbit anti-P65 (1 : 1000, Cell Signaling Technology, Danvers, MA, USA), rabbit anti-p-IκBα (1 : 1000, Cell Signaling Technology, Danvers, MA, USA), and rabbit anti-IκBα (1 : 1000, Cell Signaling Technology, Danvers, MA, USA).
7
Inflammatory Pathway Modulation Protocols
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Amlexanox was purchased from Tokyo Chemical Industry (Tokyo, Japan). STAT3 activation inhibitor SPI was purchased from BioVision (Milpitas, CA, USA). LPS from Escherichia coli O111:B4 was obtained from Sigma–Aldrich (St. Louis, MO, USA). The following primary antibodies, which were diluted at 1:1000 ratios in EveryBlot Blocking Buffer (Bio-Rad Laboratories), were used for Western blotting (WB): rabbit anti-COX2 (Cell Signaling Technology), mouse anti-iNOS (Cell Signaling Technology), rabbit anti-p-AKT (Ser473, Cell Signaling Technology), rabbit anti-AKT (Cell Signaling Technology), rabbit anti- IκBα (Cell Signaling Technology), rabbit anti-p- IκBα (Ser32, Cell Signaling Technology), rabbit anti-IKKε (Cell Signaling Technology), rabbit anti-ERK (Cell Signaling Technology), rabbit anti-p-ERK (Thr202/Tyr204, Cell Signaling Technology), rabbit anti-STAT3 (Cell Signaling Technology), rabbit anti-p-STAT3 (Tyr705, Cell Signaling Technology), rabbit anti-NF-κB p65 (Cell Signaling Technology), rabbit anti-p-NF-κB p65 (Ser536, Cell Signaling Technology), rabbit anti-JNK, rabbit anti-p-JNK (Thr183/Tyr185, Cell Signaling Technology), rabbit anti-p38 (Cell Signaling Technology), and rabbit anti-p-p38 (Thr180/Tyr182, Cell Signaling Technology), and mouse anti-β-actin (Santa Cruz Biotechnology). Other chemicals for Western blotting were obtained from Bio-Rad Laboratories.
8
Western Blot Analysis of Signaling Proteins
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Western blotting was carried out as previously described [27 (link)]. The primary antibodies used in this study were as follows: rabbit anti-Ki-67, rabbit anti-TRAF6 (Millipore), rabbit anti-TAK1, rabbit anti-phospho-TAK1, rabbit anti-IκBα, mouse anti-phospho-IκBα (Cell Signaling Technology), and mouse anti-β-actin (Boster).
9
Lipoteichoic Acid Signaling Pathway
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Lipoteichoic acid (LTA) was purchased from Sigma-Aldrich (St. Louis, MO, USA). The p38MAPK inhibitor SB203580 and NF-κB inhibitor Bay11-7082 were purchased from Calbiochem (San Diego, CA, USA). The rabbit anti-p38MAPK, rabbit anti-phospho-p38MAPK, rabbit anti-IκBα, rabbit anti-phospho-IκBα, rabbit anti-NF-κB p65 antibodies, and normal rabbit IgG were purchased from Cell Signaling (Beverly, MA, USA). The mouse anti-ACTB and rabbit NF-κB p65 antibodies were purchased from Santa Cruz Biotechnology (Dallas, TX, USA). The rabbit anti-TLR2 and rabbit anti-DEFB131 antibodies were purchased from Abcam (Cambridge, UK).
10
Immunoblotting of Cell Signaling Proteins
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Immunoblotting was performed using the following antibodies: mouse anti-cIAP1 (Santa Cruz, Heidelberg, Germany; 271418), rabbit anti-cIAP2 (Cell Signaling Technologies, Leiden, The Netherlands; 3130), rabbit anti-survivin (R&D; AF886), mouse anti-XIAP (BD, Franklin Lakes, NJ, USA; 610716), rabbit anti-IκBα (Cell Signaling Technologies; 9242), mouse anti-p-IκBα (Cell Signaling Technologies; 6249L), mouse anit-p65 (Santa Cruz; sc-8008), rabbit anti-p-p65 (Cell Signaling Technologies; 3033S), rabbit anti-NIK (Cell Signaling Technologies; 4994), mouse anti-p100 (Merck-Millipore, Darmstadt, Germany; 05-361), mouse anti-GAPDH (BioTrend, Köln, Germany; NB-29-00852), mouse anti-vinculin (Sigma/Merck; V9131), mouse anti-caspase-8 (Enzo; ADI-AAM-118-E), rabbit anti-caspase-9 (Cell Signaling Technologies; 9502S), rabbit anti-DR5 (Merck-Millipore; AB16942) and mouse anti-PARP-1 (Cell Signaling Technologies; 9546S). The following secondary horseradish peroxidase-conjugated antibodies were used: goat anti-mouse (Abcam, Berlin, Germany; ab6789) and goat anti-rabbit (Abcam; ab6721). The quantification of Western blot and the area determination of the region of interest were performed using ImageJ/FIJI [28 (link)]. Subsequent ratio calculations were performed using Microsoft Excel.
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