Rabbit anti GAPDH (2275- PC- 100) (Trevinogen; Gaithersburg, Maryland, USA); rabbit anti-mouse CEBPα, (sc-61); goat anti-human PPARγ2, (sc-22022); mouse monoclonal (E-8) anti-human PPARγ1 (sc-7273) (Santa- Cruz, Heidelberg, Germany); rabbit monoclonal anti-human phospho-p44/42 MAPK (CST 4376); rabbit anti-human p44/42 MAPK (CST-9102); rabbit monoclonal (81E11) anti-human phospho-SAPK/JNK (CST 4668); rabbit anti-human SAPK/JNK (CST 9252); rabbit monoclonal (44D4) anti-human IκB (CST-4812),), rabbit monoclonal (93H1) anti-human phospho-p65 (Ser536) (CST 3033), rabbit monoclonal (C22B4) anti-human NF-κB p65 (CST 4764) (Cell Signaling, Frankfurt, Germany); rabbit anti-mouse KLF5 (07–1580, Millipore Merck, Darmstadt, Germany); rabbit anti-mouse Cox2 (Cayman 160126; Tallinn, Estonia) and mouse monoclonal (AC-15) anti-β- Actin, (A5441, Sigma).
Anti β actin ac 15
Manufactured by Merck Group
Sourced in United States, Germany, Japan, United Kingdom, Estonia
Anti-β-actin (AC-15) is a monoclonal antibody that specifically targets the beta-actin protein. Beta-actin is a highly conserved and abundant cytoskeletal protein found in eukaryotic cells. The antibody can be used as a control or reference marker in various laboratory techniques, such as Western blotting, immunohistochemistry, and immunocytochemistry, to detect and quantify the expression of beta-actin.
Automatically generated - may contain errors
Lab products found in correlation
Anti flag m2, by Merck Group (7 mentions)
Pvdf membrane, by Merck Group (6 mentions)
Complete mini, by Roche (4 mentions)
Polyacrylamide gel, by Fujifilm (4 mentions)
Hybond, by Cytiva (3 mentions)
Phosstop, by Roche (3 mentions)
Isopropyl β d 1 thiogalactopyranoside (iptg), by Fujifilm (3 mentions)
Protease inhibitor cocktail, by Roche (3 mentions)
Sds sample buffer, by Nacalai Tesque (3 mentions)
Peroxidase conjugated anti mouse igg, by Agilent Technologies (3 mentions)
Sodium orthovanadate, by New England Biolabs (2 mentions)
Laemmli sample buffer, by Bio-Rad (2 mentions)
Nupage sample reducing agent, by Thermo Fisher Scientific (2 mentions)
Protein assay, by Bio-Rad (2 mentions)
Pvdf membrane, by Bio-Rad (2 mentions)
Las 3000 imaging system, by Fujifilm (2 mentions)
Immobilon western chemiluminescent hrp substrate, by Merck Group (2 mentions)
Anti phospho p44 42 mapk erk1 2 thr202 tyr204, by Cell Signaling Technology (2 mentions)
Bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Dulbecco s pbs, by Thermo Fisher Scientific (2 mentions)
112 protocols using anti β actin ac 15
1
Western Blot Analysis of Protein Targets
2
Syk Protein Detection in Neutrophils
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Neutrophils were lysed in Pierce radioimmunoprecipitation assay (RIPA) buffer (Thermo Fisher Scientific) containing protease inhibitors (cOmplete Mini; Roche Diagnostics, Indianapolis, IN) and sodium orthovanadate (New England BioLabs, Ipswich, MA). Proteins were resolved by SDS-PAGE under reduced conditions and transferred onto a polyvinylidene difluoride (PVDF) membrane. Membranes were blocked in Tris-buffered saline–1% Tween (TBS-T)–5% nonfat milk. Total Syk protein was detected with rabbit MAb D3Z1E anti-Syk (Cell Signaling, Danvers, MA) (1:1,000). The blots were subsequently reacted with mouse MAb AC-15 anti-β-actin (Sigma, St, Louis, MO) (1:200,000) to verify equivalent loading levels.
3
Western Blot Analysis of NCF1 Protein
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Neutrophils were lysed in reducing agent (NuPAGE® Sample Reducing Agent, ThermoFisher) and sodium dodecyl sulfate (SDS) sample buffer (4× Laemmli Sample Buffer, BioRad). Proteins were resolved by SDS-polyacrylamide gel electrophoresis under reduced conditions and transferred onto a polyvinylidene difluoride membrane. Membranes were blocked in PBS–1% Tween with 5% nonfat milk. Total NCF1 protein was detected with p47 phox (NCF1) D-10 (1:250; Santa Cruz Biotechnology, Dallas, TX). The blots were subsequently reacted with mouse monoclonal antibody AC-15 anti–β-actin (1:200,000; Sigma, St. Louis, MO).
4
Analyzing Neutrophil Signaling Pathways
5
Western Blot Protein Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Proteins were extracted with radioimmuno-precipitation assay lysis buffer (50 mM Tris/HCl, pH 7.4, 150 mM NaCl, 0.1% SDS, 1 % Nonidet P-40, 0.5 % Na deoxycholate, 1 mM EDTA), protease inhibitor (30 μg/mL) (Roche Diagnostics, Meylan, France). After sonication, protein concentrations were determined using the BioRad Protein assay (Biorad, Marnes-la-Coquette, France) according to the manufacturer's recommendations. Each sample was subjected to SDS-PAGE and then transferred onto Hybond TM-P polyvinylidene difluoride membranes (GE Healthcare, Amersham, England) which were blocked with 5% nonfat milk overnight at 4°C under constant shaking. Membranes were then incubated with primary antibodies (see below), washed and incubated with anti-mouse or anti-rabbit immunoglobulin antibodies conjugated with horseradish-peroxydase. The following primary antibodies were used for this study: 2E3F8 anti-E6HPV16 (Euromedex, Souffelweyersheim, France); DO7 anti-p53 (BD Biosciences, Le Pont de Claix, France); ABE135 anti-Sp1 (Millipore, Saint-Quentin-en-Yvelines, France) and AC15 anti-βactin (Sigma-Aldrich) antibodies. The immune complexes were revealed by chemiluminescence with Pierce ECL2 Western blotting Substrate (Thermo Scientific) using ChemiDoc XRS+ with image lab software (Biorad).
6
Western Blot Analysis of Apoptosis Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Whole-cell lysates were generated by the addition of 2× Laemmli buffer containing 10% β-mercaptoethanol to the cell pellets. Cell lysates were separated by SDS-PAGE and transferred to polyvinylidene difluoride (PVDF) membranes (Bio-Rad). Membranes were blotted with the following antibodies: anti-caspase-8 (1C12; Cell Signaling), anti-caspase-3 (8G10; Cell Signaling), anti-PCNA (D3H8P; Cell Signaling), and anti-β-actin (AC-15; Sigma-Aldrich). Peroxidase-conjugated secondary antibodies were used (BioLegend). Membranes were developed using enhanced chemiluminescence (ECL) (Thermo Scientific) and detected using a Nikon camera as previously described (82 (link)). Quantification analysis of blots was performed using ImageJ, and the β-actin signal was used as a control for normalization. Values are expressed as a percentage of the normalized signal from the mock-infected group.
7
Western Blot Analysis of Cell and Tissue Lysates
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Western blot analysis of cell lysates and skin tissue lysates was performed as previously described (Roh et al. 2021 (link)) using the following antibodies: anti-COL1A1 (3G3) (Santa Cruz Biotechnology), anti-phospho-Smad2/3 (D27F4) (#8828), anti-Smad2/3 (D7G7) (#8685), anti-phospho-AKT (ser473) (#4060), anti-phospho-AKT (thr308) (#2965), anti-AKT (pan) (#4691), anti-GAPDH (#2118) (Cell Signaling Technology), anti-β-actin (AC-15) (Sigma-Aldrich), and anti-α-SMA (MAB1420) (R&D Systems).
8
Protein Expression Analysis Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Snap-frozen cells were homogenized in lysis buffer (50 mM TRIS-HCl pH 7.6, 250 mM NaCl, 5 mM EDTA, 0.1% Triton X-100) containing complete protease inhibitor cocktail and PhosSTOP phosphatase inhibitor (Roche, Mannheim, Germany) by sonication. Immunoblotting was performed as previously described [20] (link). Antibodies were used at 1:1.000 dilution, except antibodies against p-AKT and p-RPS6 (1:2.000); β-Actin (1:10.000). Luminescent signals were detected with the digital gel documentation system Stella3200 (Raytest, Straubenhardt, Germany) and quantified using ImageJ 152-win-java8 software (National Institutes of Health, USA). Antibodies to detect PARP1 (46D11, #9532), MCL-1 (D35A5, #5453), BIM (#2819), AKT (C67E7, #4691), p-AKT (S473, D9E, #4060), ERK1/2 (137F5, #4695), p-ERK1/2 (T202/204, #4370), PRAS40 (D23C7, #2691), p-PRAS40 (T246, #2997), RPS6 (5G10, #2217), p-RPS6 (S235/236, #4858) were from Cell Signaling (Danvers, MA, USA). Anti-β-Actin (AC-15) was from Sigma-Aldrich (München, Germany).
9
Western Blot Analysis of Lamin A/C
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were washed with 1x PBS and homogenized in RIPA lysis buffer containing 100 mM Tris pH 7.4, 150 mM NaCl, 10 mM EDTA pH 8, 1% sodium deoxycholate, 1% Triton X-100 and 0.1% SDS, supplemented with protease inhibitor cocktail (cOmplete, EDTA-free, Roche), and phosphatase inhibitors (PhosSTOP, Roche). Protein concentration was determined with the Pierce BCA Protein Assay Kit and 30 µg per lane were loaded onto 8% SDS-polyacrylamide gels. Gels were then transferred to nitrocellulose membranes, blocked with 5% nonfat dry milk in TBS-T buffer (20 mM Tris pH 7.4, 150 mM NaCl and 0.05% Tween 20) and incubated overnight at 4 ºC with primary antibodies: 1:500 mouse monoclonal anti-lamin A/C (MANLAC1, provided by G. Morris) for experiments involving mouse cells, 1:1,000 rabbit polyclonal anti-lamin A/C (sc-20681, Santa Cruz Biotechnology) for experiments involving human cells or 1:10,000 anti-β-actin (AC-15, Sigma) as an endogenous control. Finally, blots were incubated with 1:10,000 goat anti-mouse (Jackson ImmunoResearch) or 1:3,000 goat anti-rabbit horseradish peroxidase (HRP) (Cell Signalling) in 1.5% nonfat dry milk in TBS-T and washed with TBS-T. Immunoreactive bands were developed with Immobilon Western chemiluminescent HRP substrate (Millipore) in a LAS-3000 Imaging System (Fujifilm). Bands were quantified using ImageJ.
10
Antibody Immunoblotting Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Monoclonal anti-SREBP-1 (2A4), anti-SCAP (9D5), and anti-HSP90 (F-8) antibodies were purchased from Santa Cruz (Dallas, TX, USA). Monoclonal anti-FLAG (M2) and anti-β-actin (AC-15) antibodies were purchased from Sigma (St Louis, MO, USA). Monoclonal anti-HA (16B12) antibody was purchased from BioLegend (San Diego, CA, USA). The polyclonal anti-SREBP-2 (RS004) antibody has been previously described46 (link). Peroxidase-conjugated affinity-purified donkey anti-mouse IgGs and peroxidase-conjugated affinity-purified donkey anti-rabbit IgGs were purchased from Jackson ImmunoResearch Laboratories (West Grove, PA, USA).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!