Fibroblast cells (healthy or asthmatic) were collected and washed with PBS, after which the proteins were extracted using the laemmli or RIPA lysis buffer (Sigma-Aldrich, Germany). All protein extracts were quantified using Bradford Protein Assay Kit, according to the manufacturer’s instructions (Bio-Rad, United States). 10 μg (for fibroblasts) of protein was separated on SDS-PAGE and transferred to a nitrocellulose membrane. Expressions of β-catenin and β-actin were assessed using rabbit anti-human CTNNB1 (Cell Signaling, United States) and mouse anti-human β-actin (A5441, Sigma, Germany), respectively. Anti-rabbit and anti-mouse IgG HRP-linked antibodies (Cell Signaling, United States) were used along with Clarity Western ECL Substrate (Bio-Rad, United States) for chemiluminescent detection of protein bands. Western blot analysis of cell fractions from asthmatic bronchial fibroblast using Cell Fractionation Antibody Sampler Kit #11843 showing cytoplasmic (C.F), organellular/membrane (M.F), and nuclear/cytoskeletal localization (N.F.). Whole-cell lysates (WCL) represent total protein. The fractionation was done under two conditions, the cultivation with high glucose medium and low glucose medium.
Anti rabbit and anti mouse igg hrp linked antibodies
Manufactured by Cell Signaling Technology
Sourced in United States
Anti-rabbit and anti-mouse IgG HRP-linked antibodies are secondary antibodies used in Western blotting and ELISA applications. These antibodies are conjugated with horseradish peroxidase (HRP), which enables the detection of target proteins through a colorimetric or chemiluminescent reaction.
Automatically generated - may contain errors
Lab products found in correlation
Sorafenib, by Cell Signaling Technology (2 mentions)
Salirasib, by Merck Group (2 mentions)
Pf573228, by Bio-Techne (2 mentions)
Gefitinib, by Selleck Chemicals (2 mentions)
Anti adam10, by Abcam (2 mentions)
Fak antibody sampler kit, by Cell Signaling Technology (2 mentions)
Phospho egf receptor antibody sampler kit, by Cell Signaling Technology (2 mentions)
Phospho erk1 2 pathway sampler kit, by Cell Signaling Technology (2 mentions)
Alpha tubulin dm1a antibody, by Cell Signaling Technology (2 mentions)
Gi254023x, by Bio-Techne (2 mentions)
Anti adam17, by Abcam (2 mentions)
U0126, by Cell Signaling Technology (2 mentions)
Ir dye coupled secondary antibodies, by LI COR (2 mentions)
Gw280264x, by Aobious (2 mentions)
Mouse anti human β actin, by Merck Group (1 mentions)
Ripa lysis buffer, by Merck Group (1 mentions)
Bradford protein assay kit, by Bio-Rad (1 mentions)
Clarity western ecl substrate, by Bio-Rad (1 mentions)
Lps from escherichia coli 0111 b4, by Merck Group (1 mentions)
Recombinant human saa, by Thermo Fisher Scientific (1 mentions)
9 protocols using anti rabbit and anti mouse igg hrp linked antibodies
1
Fibroblast β-catenin Expression Analysis
2
Epigenetic Chromatin Modification Enzymes PCR Array
3
Characterization of EGFR Signaling Inhibitors
4
CoCl2-Induced Hypoxia Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
CoCl2 was purchased from Sigma-Aldrich (St. Louis, MO, USA). Primers for GAPDH, GPC3, and HIF-1α were synthesized by Sangon Biotech (Shanghai, China). The protease inhibitor was purchased from Roche (Mannheim, Germany). PowerUp™ SYBR™ Green Master Mix was purchased from Applied Biosystems (Foster City, CA, USA) Mouse anti-human monoclonal antibodies against β-actin and GPC3 were acquired from Santa Cruz Biotechnology (1:1000, Santa Cruz, CA, USA). Rabbit anti-human monoclonal antibodies against HIF-1α, c-myc, sp1, PARP and caspase-3 were obtained from Cell Signaling Technology (1:1000, Danvers, MA, USA). Anti-rabbit and anti-mouse IgG HRP-linked antibodies were procured from Cell Signaling Technology (1:2000, Danvers, MA, USA). RIPA lysis buffer was obtained from Beyotime Institute of Biotechnology (Shanghai, China).
5
Characterization of EGFR Signaling Inhibitors
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Salirasib (5 μM, #SML1166; Sigma), Sorafenib (5 μM, #8705) and U0126 (20 μM, #9903; Cell Signaling Technology), Gefitinib (10 μM, Iressa, #S1025; Selleck-Chem), GI254023X (20 μM, #3995) and PF573228 (10 μM, #3239; Tocris), and GW280264X (20 μM, AOB3632; Aobious) were all used at the indicated concentrations. Monoclonal anti-B5 (VMC-20) was a kind gift of Gary H. Cohen and Roselyn J Eisenberg (University of Pennsylvania)40 (link). Phospho-EGF receptor antibody sampler kit (#9922, containing #4267, 3777, 2237), phospho-Erk1/2 pathway sampler kit (#9911, containing #9427, 9154, 4370, 11989), FAK antibody sampler kit (#9330, containing #3281, 3284, 8556, 13009), alpha-tubulin DM1A antibody (#3873), and anti-rabbit and anti-mouse IgG HRP-linked antibodies (#7074, 7076) were purchased from Cell Signaling Technology and used at 1:1000. Anti-ADAM10 (#ab124695) and anti-ADAM17 (#ab2051) were purchased from Abcam and used at 1:1000. IRDye-coupled secondary antibodies were purchased from Licor and used at 1:10'000. Alexa Fluor-conjugated secondary antibodies and phalloidin were purchased from Invitrogen/Thermo Fischer Scientific and used at 1:1000 or 1:100, respectively.
6
Inflammatory Signaling Pathway Analyses
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Antibodies and other reagents were obtained from the following suppliers: anti-MCP-1 (Abcam, Cambridge, UK); anti-β-actin (C4) (Santa Cruz Biotechnology, Inc., Dallas, TX, USA); anti-phospho-extracellular signal-regulated kinase (ERK) (Thr202/Tyr204), anti-phospho-p38 (Thr180/Tyr182), anti-phospho-c-Jun N-terminal kinase (JNK) (Thr183/Tyr185), anti-rabbit and anti-mouse IgG-HRP-linked antibodies (Cell Signaling Technology, Inc., Danvers, MA, USA); rifampicin, Protease Inhibitor Cocktail (EDTA free) (100x), and Phosphatase Inhibitor Cocktail (all from Nacalai Tesque Inc., Kyoto, Japan); Microcystin-LR, penicillin-streptomycin solution (x100), Dulbecco’s modified Eagle's medium (DMEM) supplemented with low glucose, and SP600125 (all from Wako Pure Chemical Industries Ltd., Osaka, Japan); probenecid (Sigma-Aldrich, St Louis, MO, USA), and fetal bovine serum (FBS; Biowest S.A.S., Nuaillé, France).
7
Western Blot Analysis Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Lysates were run on 10% Bis-Tris Plus polyacrylamide gels (Thermo Fisher) and transferred to PVDF membrane (Bio-Rad). Membranes were probed with the indicated antibodies and were developed by Western Lightning Plus ECL (Perkin Elmer) and imaged on a ChemiDoc MP Imaging System (Bio-Rad). Antibodies, rabbit anti-Flag (cat# F7425), and mouse anti-β-tubulin (cat# T8328) were purchased from Sigma-Aldrich. Rabbit anti-HA (cat# 71-5500) and mouse anti-GAPDH (GA1R) (cat# MA5-15738) were purchased from Invitrogen. Anti-rabbit and anti-mouse IgG, HRP-linked antibodies were purchased from Cell Signaling (cat# 7074 and 7076).
8
Synergistic Effects of MEK and PARP Inhibition
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The MEK1/2 inhibitor pimasertib and hypoxia-activated prodrug evofosfamide were provided by EMD Serono Research and Development Institute (Billerica, MA) and Threshold Pharmaceuticals (South San Francisco, CA). Both reagents were dissolved in DMSO to make a 10mM stock solution and were stored at −20°C. The PARP inhibitors olaparib and rucaparib were a gift from the University College Hospital MacMillan Cancer Center (London, UK). The following reagents were used: Thiazolyl Blue Tetrazolium Bromide (MTT); for immunoblotting, the following antibodies were used: anti–β-actin and anti-calnexin as loading controls; anti-cleaved-PARP, anti-p-ERK, anti-ERK, anti-γH2AX (Cell Signaling Technology) and secondary antibodies anti-mouse and anti-rabbit IgG HRP linked antibodies (Cell Signaling Technology). For Immunofluorescence analysis, anti-BRCA2 (Santa Cruz Biotechnology), anti-RAD51 (Abcam) and anti-phosphorylated histone-H2AX (Merck Millipore) antibodies were used.
9
Investigating Cell Apoptosis in PC12 Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Fluorescent probe 2,7-dichloro uorescein diacetate (DCF-DA) and (4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium (MTT) were purchased from Sigma. RPMI 1640 was purchased from Bioidea Company and FBS was purchased from Gibco. Rabbit monoclonal anti-serums against procaspase-3, caspase-3cleaved, Bcl-2, caspase-8, and ß-actin, rabbit polyclonal anti-serum against Bax, and anti-mouse and antirabbit IgG HRP-linked antibodies were purchased from Cell signaling. Cadmium and minocycline were purchased from Tinab Shimi. Polyvinylidene uoride (PVDF) membrane was purchased from Bio-Rad.
Cell culture PC12 (pheochromocytoma cell line of rat) cells were obtained from the Pasteur Institute of Iran (Tehran). Cells were cultured in RPMI-1640 medium and incubated at 37˚C with 5% CO 2 . The medium contained 10% fetal bovine serum and 5% antibiotic mixture (100 U/mL penicillin and 100 µg/mL streptomycin) and was changed every 2-3 days.
Cell culture PC12 (pheochromocytoma cell line of rat) cells were obtained from the Pasteur Institute of Iran (Tehran). Cells were cultured in RPMI-1640 medium and incubated at 37˚C with 5% CO 2 . The medium contained 10% fetal bovine serum and 5% antibiotic mixture (100 U/mL penicillin and 100 µg/mL streptomycin) and was changed every 2-3 days.
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!