Rabbit polyclonal NP and NS1 antibodies were produced in our lab [35 (link)]. The commercial antibodies used are as follows. Goat anti-porcine IL-1β antibody (BAF681): R&D Systems (Minneapolis, MN, USA); rabbit anti-porcine caspase-1 (p20) antibody (PAB592Po01): Cloud-Clone Corp. (Houston, TX, USA); mouse anti-Myc-tag antibody (#2276), mouse anti-β-actin antibody (#3700), rabbit anti-DRP1 antibody (#8570) and rabbit anti-phospho-DRP1 (S616) antibody (#3455): Cell Signaling Technology (Beverly, MA, USA); mouse anti-FLAG M2 antibody (F3165): Sigma; IRDye 680RD donkey anti-rabbit (926-68073), IRDye 800CW donkey anti-mouse (926-32212) and IRDye 800CW donkey anti-goat (926-32214) antibodies: LI-COR Biosciences (Lincoln, NE, USA). The following reagents were used: lipopolysaccharide (LPS) (L3024) and N-acetyl L-cysteine (NAC) (A9165): Sigma; Necrostatin-1 (Nec-1) (BML-AP309): Enzo Life Sciences (Farmingdale, NY USA); Mdivi-1 (ab144589): Abcam (Cambridge, MA, USA).
Mouse anti β actin antibody
Manufactured by Cell Signaling Technology
Sourced in United States
Mouse anti-β-actin antibody is a laboratory reagent used to detect and quantify the presence of the β-actin protein in biological samples. β-actin is a commonly used reference or housekeeping protein in various cell and molecular biology experiments.
Automatically generated - may contain errors
Lab products found in correlation
Ripa buffer, by Thermo Fisher Scientific (4 mentions)
Pvdf membrane, by Merck Group (3 mentions)
Hrp conjugated goat anti rabbit igg antibody, by Cell Signaling Technology (2 mentions)
Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (2 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (2 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions)
Mouse anti flag m2 antibody, by Merck Group (1 mentions)
Mouse anti myc tag antibody, by Cell Signaling Technology (1 mentions)
Rabbit anti drp1 antibody, by Cell Signaling Technology (1 mentions)
Irdye 800cw donkey anti goat 926 32214, by LI COR (1 mentions)
Mdivi 1 ab144589, by Abcam (1 mentions)
Rabbit anti phospho drp1 s616 antibody, by Cell Signaling Technology (1 mentions)
Irdye 680rd donkey anti rabbit 926 68073, by LI COR (1 mentions)
Necrostatin 1 nec 1 bml ap309, by Enzo Life Sciences (1 mentions)
Mouse anti flag antibody, by Merck Group (1 mentions)
Anti v5, by Merck Group (1 mentions)
Supersignal west pico detection kit, by Thermo Fisher Scientific (1 mentions)
Horseradish peroxidase conjugated secondary antibody, by Santa Cruz Biotechnology (1 mentions)
Rabbit anti phospho p44 42 mapk erk1 2 antibody, by Cell Signaling Technology (1 mentions)
33 protocols using mouse anti β actin antibody
1
Antibodies and Reagents for Porcine Cell Study
2
Western Blot Analysis of Recombinant hOAS1
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hOAS1 proteins expressed in bacteria or in mammalian cells were separated on a 10% SDS-PAGE gel and transferred to a nitrocellulose membrane at 100 V for 1 h. The membrane was incubated in blocking buffer (1 X Tris-buffered saline containing 5% non-fat dry milk and 0.1% Tween 20) at room temperature for 1 h and then cut into strips. To detect the hOAS1 isoform proteins expressed in bacteria, the membrane strips were incubated with anti-V5 (Sigma-Aldrich, St. Louis, MO, USA; 1:1000) antibody at 4 °C overnight. To detect overexpressed hOAS1 isoforms in HEK293 cell lysates, the membrane strips were incubated at 4 °C overnight with mouse anti-Flag antibody (Sigma-Aldrich, St. Louis, MO, USA; 1:1000) or mouse anti-β actin antibody (Cell signaling, Danvers, MA, USA; 1: 10,000). The membranes were washed three times for 10 min with 1 X Tris-buffered saline containing 0.1% Tween 20, and then incubated with a horseradish peroxidase-conjugated secondary antibody (Santa Cruz Biotechnology, Dallas, TX, USA) for 1 h at room temperature. After washing, the membrane strips were processed for enhanced chemiluminescence using a Super-Signal West Pico detection kit (Thermo Fisher Scientific, Waltham, MA, USA) according to the manufacturer’s protocol.
3
Immunoblot Analysis of MAPK Signaling
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The following commercial antibodies from Cell Signaling Technology were used : rabbit anti-p38 MAPK antibody (catalog no. 8690, 1:4000 dilution); rabbit anti-phospho-p38 MAPK antibody (catalog no. 4511, 1:2000 dilution); mouse anti-p44/42 MAPK (Erk1/2) antibody (catalog no. 4696, 1:2000 dilution); rabbit anti-phospho-p44/42 MAPK (Erk1/2) antibody (catalog no. 4370, 1:4000 dilution); mouse anti-β-actin antibody (catalog no. 3700, 1:4000 dilution); and peroxidase-conjugated goat anti-rabbit IgG (catalog no. 7074, 1:4000 dilution). Peroxidase-conjugated sheep anti-mouse IgG antibody was obtained from Cytiva (Tokyo, Japan; catalog no. NA931, 1:2000 dilution). Peroxidase-conjugated donkey anti-rat IgG antibody was obtained from Jackson ImmunoResearch (West Grove, PA, USA; catalog no. 712-035-150, 1:5000 dilution). Rat anti-mCherry antibody was obtained from Thermo Fisher Scientific (catalog no. M11217, 1:1000 dilution).
4
Investigating SKOV-3 Ovarian Cancer Cells
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Human ovarian adenocarcinoma cells (SKOV-3) were purchased from American Type Culture Collection (Manassas, VA, USA). RES was purchased from MilliporeSigma (Burlington, MA, USA). MTT reagent (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) was purchased from MilliporeSigma. Guava® Cell Cycle Reagent was purchased from Luminex Corporate (Austin, TX, USA). Annexin V was purchased from ImmunoTools (Friesoythe, Niedersachsen, Germany). Propidium iodide was purchased from MilliporeSigma. Rabbit anti-phospho p38 antibody, rabbit anti p38 antibody, rabbit anti-phospho ERK1/2 antibody, mouse anti-ERK1/2 antibody, rabbit anti-phospho JNK antibody, rabbit anti-JNK antibody, rabbit anti-phospho AKT (Ser473) antibody, mouse anti-AKT antibody, rabbit anti-cyclin A2 antibody, rabbit anti-cyclin B1 antibody, rabbit anti-cyclin E1 antibody, rabbit anti-caspase-9 antibody, rabbit anti-caspase-3 antibody, rabbit anti-PARP antibody, mouse anti-β-actin antibody, and DAPI (4, 6-diamidino-2-phenylindole, dihydrochloride) were purchased from Cell Signaling Technology (Danvers, MA, USA). Goat anti-mouse IgG-IRDye®800CW and goat anti-rabbit IgG-IRDye®680RT were purchased from Li-COR Biosciences (Lincoln, NE, USA). Goat anti-rabbit conjugated with Alexa488 and Goat anti-mouse conjugated with Alexa594 were purchased from Thermo Fisher Scientific (Waltham, MA, USA).
5
Quantification of β-catenin in Cardiomyocytes
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Whole-cell lysates were generated by scraping cultures of matured cardiomyocytes treated with CHIR99021, N-cadherin antibody, or combination for 1 day and resuspended in RIPA lysis buffer (1% Nonidet P-40, 0.5% deoxycholate, 5 M NaCl, and 1 M Tris pH 7.4) for 10 min at 4 °C and then centrifuged at 11,400g for 20 min at 4 °C for the isolation of the cytosolic fractions. Quantification of protein concentration was determined using Pierce BCA Protein Assay kit (Thermo-scientific). Protein lysate of 60 μg per sample was loaded onto 4–20% Mini-PROTEAN TGX Stain-Free Precast Gels (Bio-Rad), resolved by electrophoresis, and transferred to nitrocellulose membrane (Bio-Rad). Membranes were blocked with 5% skimmed milk in TBS + 1% Tween-20 and probed overnight with primary antibodies. The antibodies were rabbit anti-β-catenin antibody (1:2500; Abcam ab32572) and mouse anti-β-actin antibody (1:1000; Cell Signaling). Membranes were washed and incubated for 1 h with their respective IgG-HRP-conjugated secondary antibody (Santa Cruz) in 5% skimmed milk in TBS + 1% Tween-20 and developed using ECL substrate (Bio-Rad).
6
Western Blot Analysis of TMEM16A
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Cells were washed twice with ice-cold phosphate-buffered saline (PBS) and dissolved in lysis buffer (150 mM NaCl, 20 mM Tris, 5 mM EDTA, pH 7.5, 1% Triton X-100 and supplemented with 1 mM PMSF). After protein quantification, samples containing 100 µg proteins were denatured and electrophoresed using 10% SDS-PAGE and then transferred onto PVDF membranes. After blocking with 5% (w/v) nonfat milk and washing with Tris-buffered saline–Tween solution (TBST), the membranes were incubated overnight at 4°C with primary anti-TMEM16A polyclonal antibody (Abcam, ab53212; 1∶100) and mouse anti-β-actin antibody (Cell Signaling; 1∶500), respectively. All other primary antibodies were purchased from Cell Signaling Technology. After washing, the blots were incubated with secondary antibody conjugated to horseradish peroxidase (Sigma; 1∶5,000) for 1 h at room temperature and visualized using an enhanced chemiluminescence kit (Amersham, Little Chalfont, UK) on X-ray film (Millipore Corporation, Billerica, USA).
7
Western Blot Analysis of Cellular Proteins
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Whole cell extracts from cell lines were obtained using RIPA buffer (Thermofisher) containing a protease inhibitor cocktail (Roche) and benzonase (Thermofisher). Protein concentrations were quantified using a BCA gold assay (Thermofisher). Equal amounts of total protein were prepared in 4X NuPAGE LDS Sample Buffer (NP0007, Invitrogen) and 5% BME, and boiled at 95°C for 5 min. The protein samples were resolved on 4–12% NuPAGE Bis-Tris gel with protein molecular weight standards (PageRuler Prestained Protein Ladder, 10 to 180 kDa, Thermofisher). The gels were transferred onto PVDF membranes using the Trans-Blot Turbo Transfer System (Bio-Rad). Primary antibodies used in this study included: rabbit anti-c-Myc antibody [Y69] (ab32072, Abcam, 1:1000), mouse anti-β-Actin antibody (3700, Cell Signaling, 1:5000). Fluorescent secondary antibodies used in this study were IRDye 680RD Donkey anti-Mouse IgG Secondary Antibody and IRDye 800CW Donkey anti-Rabbit IgG Secondary Antibody (Licor, 1:5000).
8
Quantitative Western Blot Analysis of Autophagy Markers
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The cellular total protein were obtained by a protein extraction kit. Besides, the Bradford protein assay kit (BESTBIO) was used to determine the protein concentration. 30 μg or 50 μg of total cell extracts were analyzed by Western blotting. The proteins were loaded onto 15% SDS-PAGE gels and transferred onto a polyvinylidene difluoride (PVDF) membrane (Millipore, USA). Membranes were incubated with 1 μg/ml Rabbit anti-ATG4a antibody (Polyclonal, ABGENT), 2 μg/ml mouse anti-SQSTM1/p62 antibody (Monoclonal, Abcam), 2 μg/ml rabbit anti-LC3 antibody (Cell Signaling Technology, Inc.), 2.5 μg/ml rabbit anti-GAPDH antibody (Monoclonal, Cell Signaling Technology, Inc.) or 2 μg/ml mouse anti-β-actin antibody (Monoclonal, Cell Signaling Technology, Inc.). After that, the membranes were incubated with 1 μg/ml HRP-conjugated goat anti-rabbit IgG secondary antibody or 1 μg/ml HRP-conjugated goat anti-mouse IgG secondary antibody (Proteintech Group, Inc.). Immunoreactive band analysis was performed by using the enhanced western bright ECL reagent (Advansta, United States). Densitometry analyses of western blots were performed with ImageJ software (version 1.46). Western blots were developed to be linear in the range used for densitometry. All results were expressed as a relative ratio to the β-actin or GAPHD. At least three or four independent western blot experiments were performed.
9
Zerumbone Attenuates LPS-Induced Inflammation
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Zerumbone (ZER) was purchased from Sigma Aldrich (No.Z3902, St. Louis, MO, USA), dissolved in DMSO (Dimethyl sulfoxide). Bifendate (BIF) was purchased from Beijing Yuehe Pharmaceutical Factory (H11020980, Beijing, China). LPS was purchased from Beyotime Biotechnology (No.S1732, Shanghai, China). RPMI 1640 Medium was purchased from Hyclone (Logan, UT, USA). Fetal bovine serum (FBS) was purchased from Gibco (Gaithersburg, MA, USA). The assay kits for ALT, AST, SOD, GSH-Px, GSH, and MDA were purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing, China). Bicinchoninic acid (BCA) assay kit and MTT were purchased from Solarbio (Beijing, China). Mouse IL-6 and TNF-α ELISA kits was purchased from ShangHai Haling Biological Technology Co., Ltd. (ShangHai, China). Rabbit anti-TLR4 antibody (#14358, 1:1000), rabbit anti-COX-2 antibody (#4842, 1:1000), rabbit anti-p-NF-κB p65 (Ser536) antibody (#3033, 1:1000), mouse anti-β-Actin antibody (#3700, 1:2000) were purchased from Cell Signaling Technology (Danvers, MA, USA). All other chemicals and reagents were purchased from local firms.
10
Apoptosis Induction and Analysis
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Fetal bovine serum (FBS), penicillin-streptomycin, and the cell culture medium (RPMI 1640) were prepared from Gibco BRL, UK. The culture plates were provided by SPL (Life Sciences, Korea). Annexin V-FITC/PI kit and PI were obtained from BD Pharmingen™. Ethidium bromide, acridine orange, 3-(4,5-dimethylthiazole)-2,5-diphenyltetrazolium bromide (MTT), and Ac-DEVD-p-nitroaniline (pNA) as a chromogenic substrate of caspase-3 were prepared from Sigma-Aldrich. Caspase-9 and caspase-8 antibodies were provided by Abcam, and mouse anti-β-actin antibody was obtained from cell signaling. The secondary antirabbit and antimouse HRP-conjugate antibodies were prepared from Razi BioTech (Tehran, Iran). All other chemicals were provided by Merck (Darmstadt, Germany) and Sigma-Aldrich.
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