Anti-Lyn(Santa Cruz, CA, sc-15), anti-Vimentin (Santa Cruz,sc-6260), anti-Smad2/3(Santa Cruz, CA, sc-8332), anti-p-Smad2/3(Santa Cruz, CA, sc-11,769), and anti-β-actin(Santa Cruz, CA, sc-130,656) were purchased from Santa Cruz Biotechnology. Anti-E-cadherin (Abcam,ab11512), and anti-α-SMA (Abcom,ab5694) were purchased from Abcam Biotechnology. Anti-p-Lyn (CST, #2731) was purchased from Cell Signaling Technology. Nonsilencing siRNA control, Lyn-specific siRNA were purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Recombinant lentivirus containing Lyn or a GFP-only virus control was obtained from Cyagen (Shanghai, China).
Anti smad2 3
Manufactured by Santa Cruz Biotechnology
Sourced in United States, United Kingdom
Anti-Smad2/3 is a laboratory reagent used in research applications. It functions as an antibody that specifically binds to the Smad2 and Smad3 proteins, which are intracellular signal transducer proteins involved in the transforming growth factor-beta (TGF-β) signaling pathway.
Automatically generated - may contain errors
Lab products found in correlation
Anti p smad2 3, by Santa Cruz Biotechnology (4 mentions)
Anti vimentin, by Santa Cruz Biotechnology (3 mentions)
Anti β actin, by Santa Cruz Biotechnology (2 mentions)
Anti e cadherin, by Abcam (1 mentions)
Anti lyn, by Santa Cruz Biotechnology (1 mentions)
Image pro plus 6, by Media Cybernetics (1 mentions)
Anti tgf β1, by Abcam (1 mentions)
Hiseq platform, by Illumina (1 mentions)
Anti gfp antibody, by Thermo Fisher Scientific (1 mentions)
Anti tau1, by Merck Group (1 mentions)
Anti map2, by Merck Group (1 mentions)
Anti sara, by Santa Cruz Biotechnology (1 mentions)
Anti gadd34, by Santa Cruz Biotechnology (1 mentions)
Anti tβri, by Santa Cruz Biotechnology (1 mentions)
Horseradish peroxidase conjugated secondary antibody, by GE Healthcare (1 mentions)
Mini protean tetra cell, by Bio-Rad (1 mentions)
Luminol reagent, by Santa Cruz Biotechnology (1 mentions)
Anti erk1 2, by Santa Cruz Biotechnology (1 mentions)
Anti p erk1 2, by Santa Cruz Biotechnology (1 mentions)
Anti phospho fak y397, by Abcam (1 mentions)
15 protocols using anti smad2 3
1
Lyn Signaling in Epithelial-Mesenchymal Transition
2
Histological Analysis of Kidney Fibrosis
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Kidneys were fixed with 4% paraformaldehyde for at least 24 hours, and then they were dehydrated, embedded and cut into 5‐μm sections. The sections were stained with haematoxylin & eosin (H&E) and Masson's trichrome for general morphological assay.
Immunohistochemical detection was conducted with general approaches. Anti‐α‐SMA (1:500, CST, Boston, MA, USA, #56856), anti‐TGF‐β1 (1:200, Abcam, Cambridge, UK, #92486) and anti‐Smad2/3 (1:200, Santa Cruz, Santa Cruz, CA, USA, #398844) were used as the primary antibodies. Image Pro Plus 6.0 (Media Cybernetics, Inc., Rockville, MD, USA) image analysis software was used to calculate the mean optical density value. At least 5 non‐overlapping cortical fields per section were analysed for each animal in a blinded manner.
Immunohistochemical detection was conducted with general approaches. Anti‐α‐SMA (1:500, CST, Boston, MA, USA, #56856), anti‐TGF‐β1 (1:200, Abcam, Cambridge, UK, #92486) and anti‐Smad2/3 (1:200, Santa Cruz, Santa Cruz, CA, USA, #398844) were used as the primary antibodies. Image Pro Plus 6.0 (Media Cybernetics, Inc., Rockville, MD, USA) image analysis software was used to calculate the mean optical density value. At least 5 non‐overlapping cortical fields per section were analysed for each animal in a blinded manner.
3
Smad2/3 ChIP-Seq in Xenopus Embryos
4
Antibody Characterization for Cellular Pathways
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The following antibodies were used in this study: anti-SARA [mouse, sc-133071; 1:200 for immunoblotting and 1:100 for immunofluorescence (IF)], anti-PP1c (mouse, sc-7482; 1:50 for IF), anti-GADD34 (mouse, sc-373815; 1:50 for IF), anti-TβRI (mouse, sc-101574; 1:100 for IF) and anti-Smad2/3 (mouse, sc-398844; 1:100 for IF); all these antibodies were purchased from Santa Cruz Biotechnology (Dallas, Texas, United States). The antibody anti-MAP2 (rabbit, 1:500) and anti-Tau-1 (mouse, 1:500) were from Merck Millipore (Darmstadt, Germany). Antibody anti-pSamd2/3 (rabbit, s465/s467, E8F3R, 1:50 for IF) was from Cell Signaling (Danvers, MA, United States) and anti-phospho TβRI (rabbit, ser165, Lot: DY1241; 1:50 for IF) was acquired from Elabscience (Houston, Texas, United Sates).
5
Protein Expression Analysis via Western Blot
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Cell lysates were resolved in 10% sodium dodecyl sulfate polyacrylamide mini gels (Mini-PROTEAN® Tetra Cell; Bio-Rad Laboratories, Inc., Hercules, CA, USA) and transferred onto Whatman® Protran® nitrocellulose membranes (GE Healthcare Bio-Sciences AB, Uppsala, Sweden). Membranes were blocked with 5% (weight/volume [w/v]) bovine serum albumin in Tris-buffered saline with Tween 20 for 1 hour, incubated with primary antibodies against target proteins, and then incubated with horseradish peroxidase-conjugated secondary antibody (GE Healthcare). The proteins were visualized using Western blotting Luminol reagent (Santa Cruz Biotechnology Inc., Dallas, TX, USA). The following antibodies were used: anti-PKN1 (sc-7161), anti-Smad2/3 (sc-6032), anti-pSmad2 (sc-135644), anti-ERK1/2 (sc-135900), anti-pERK1/2 (sc-7383), vimentin (sc-7557), E-cadherin (sc-7870), anti-FAK (sc-558), actin (sc-8432) (Santa Cruz Biotechnology), and anti-FAK (phospho Y397) (ab4803; Abcam, Cambridge, UK).
6
Flow Cytometry Analysis of T Cell Activation Markers
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Flow cytometry antibodies for anti-CD127 (A7R34) and CD122 (2-1221-82) were purchased from eBioscience (San Diego, CA, USA): anti-CD3ε (BD Biosciences, San Diego, CA, USA), anti-CD8 (4B11) (Invitrogen, Carlsbad, CA, USA), anti-KLRG1 (2F1) (Southern Biotech, Birmingham, AL, USA), anti-TGFβRI, anti-Smad2/3, anti-Smad4 (Santa Cruz Biotechnology, Santa Cruz, CA, USA), anti-TGFβRII (R&D Systems), anti-T-bet (BioLegend, San Diego, CA, USA), and anti-RGS3. The anti-RGS3 antibody has been described previously [10 (link)]. DAPI was purchased from Invitrogen. Extracellular and intracellular marker staining was performed as previously described [11 (link)]. Flow cytometric analysis was performed using an LSRII flow cytometer (Becton Dickinson, Franklin Lakes, NJ, USA). Data were analyzed using Flow Jo software (Tree Star, Ashland, OR, USA). The cells were fixed and permeabilized with methanol and formaldehyde prior to staining for analysis with the Amnis Imagestream imaging flow cytometer. Data were analyzed using Amnis IDEAS software v4.0 (Amnis Corporation, Seattle, WA, USA).
7
Protein Expression Analysis in Cardiovascular Tissues
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Total proteins, membrane proteins and cytoplasmic proteins were extracted (Beyotime,
Haimen, China) and equal amounts of proteins were run on SDS-PAGE. After electrophoresis,
the proteins were transferred onto Millipore PVDF membranes (Billerica, Massachusetts,
USA). The membranes were immersed in 5% nonfat milk and incubated with anti-Angiotensin
converting enzyme (ACE) (1:200) (Santa Cruz, Dallas, Texas, USA), anti-Angiotensin II type
1 receptor (AT1 receptor) (1:1,000) (Abcam, Cambridge, MA, USA), anti-periostin (1:200)
(Santa Cruz), anti-collagen I (1:400) (Wuhan Boster, Wuhan, China), anti-TGF-β1 (1:200)
(Santa Cruz, USA), anti-Smad2/3 (1:200) (Santa Cruz), anti-p-ERK1/2 (1:500) (Bioss,
Beijing, China), anti-ERK1/2 (1:500) (Bioss), anti-GLUT4 (1:200) (Santa Cruz), anti-p-Akt
(1:200) (Santa Cruz) and anti-Akt (1:200) (Santa Cruz, USA) primary antibodies overnight.
After washing with TBS-T buffer, the membranes were incubated for 45 min with horseradish
peroxidase (HRP)-conjugated secondary antibody (1:5,000) (Beyotime) diluted in 5% nonfat
milk. The proteins were developed using ECL reagent in the dark and quantified using
Gel-Pro Analyzer (Media Cybernetics, Inc., Rockville, MD, USA).
Haimen, China) and equal amounts of proteins were run on SDS-PAGE. After electrophoresis,
the proteins were transferred onto Millipore PVDF membranes (Billerica, Massachusetts,
USA). The membranes were immersed in 5% nonfat milk and incubated with anti-Angiotensin
converting enzyme (ACE) (1:200) (Santa Cruz, Dallas, Texas, USA), anti-Angiotensin II type
1 receptor (AT1 receptor) (1:1,000) (Abcam, Cambridge, MA, USA), anti-periostin (1:200)
(Santa Cruz), anti-collagen I (1:400) (Wuhan Boster, Wuhan, China), anti-TGF-β1 (1:200)
(Santa Cruz, USA), anti-Smad2/3 (1:200) (Santa Cruz), anti-p-ERK1/2 (1:500) (Bioss,
Beijing, China), anti-ERK1/2 (1:500) (Bioss), anti-GLUT4 (1:200) (Santa Cruz), anti-p-Akt
(1:200) (Santa Cruz) and anti-Akt (1:200) (Santa Cruz, USA) primary antibodies overnight.
After washing with TBS-T buffer, the membranes were incubated for 45 min with horseradish
peroxidase (HRP)-conjugated secondary antibody (1:5,000) (Beyotime) diluted in 5% nonfat
milk. The proteins were developed using ECL reagent in the dark and quantified using
Gel-Pro Analyzer (Media Cybernetics, Inc., Rockville, MD, USA).
8
Peri-infarct Area Protein Analysis
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Rats were anesthetized and decapitated on a given day, and brain tissues from the ipsilateral peri-infarct area were collected and extracted. Anti-ALK5 (1:1000, Abcam), anti-Smad2/3 (1:1000; Santa Cruz) and anti-pSmad2/3 (1:1000; Santa Cruz) were used as primary antibodies. After the secondary antibody reaction, the bands were visualized by electrochemiluminescence (Millipore, Darmstadt, Germany). The positive pixel area was detected by an image analysis software (BIO-RAD Gel Doc 2000, Watertown, MA, USA). Western blot quantification was performed by densitometry and was normalized to GAPDH. All experiments were conducted under the same conditions and repeated three times.
9
Malonic Acid Modulates Oxidative Stress Response
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Malonic acid (MA), 2′, 7′-dichlorodihydrofluorescein diacetate (DCF-DA), dimethyl sulfoxide, and epigallocatechin-3-gallate (EGCG), as a positive control of ROS induction, were purchased from Sigma-Aldrich (St. Louis, MO, USA). Fetal bovine serum (FBS), Dulbecco’s modified Eagle’s medium (DMEM), and trypsin-ethylenediaminetetraacetic acid were purchased from Welgene (Gyeongsan, Korea). SYBR green was purchased from GeneAll (Seoul, Korea). All antibodies (anti-heme oxygenase 1 (HO-1), anti-superoxide dismutase 1 (Sod1), anti-nuclear factor-erythroid 2-related factor-2 (Nrf2), anti-p-inhibitor of NF-κB (IκB), anti-IκB, anti-p-p65, anti-p65, anti-p50, anti-Cox-2, anti-IL-6, anti-TNF-α, anti-p-JNK, anti-JNK, anti-p-ERK, anti-ERK, anti-p-p38, anti-p38, anti-p-c-Jun, anti-c-Jun, anti-c-Fos, anti-p-Smad2/3, anti-Smad2/3, anti-collagen type I alpha 1 (Col1a1), anti-Col3a1, and anti-glyceraldehyde 3-phosphate dehydrogenase (Gapdh)) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA).
10
Immunoblotting Analysis of Cardiac Proteins
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