Western blotting according to standard procedure. The difference is that the Trans-Blot SD Semi-Dry Electrophoretic Transfer Cell (Trans-BlotR TurboTM Transfer SYSTEM, BIO-RAD, USA) was then used to transfer total protein onto polyvinylidene fluoride membranes (Roche, USA). The following primary antibodies were used: anti-EMP3 (Abcam), α-tubulin, anti-MMP2, anti-MMP9, and anti-N-cadherin from Proteintech Group, Inc. ImageJ was used to perform densitometric analyses of immunoblots.
Anti mmp9
Manufactured by Proteintech
Sourced in China, United States
Anti-MMP9 is a laboratory reagent that can be used to detect and quantify the presence of matrix metalloproteinase 9 (MMP9) in samples. MMP9 is an enzyme involved in the breakdown of extracellular matrix proteins and plays a role in various biological processes. This product can be utilized in research applications that require the identification and measurement of MMP9 levels.
Automatically generated - may contain errors
Lab products found in correlation
Anti mmp2, by Proteintech (21 mentions)
Anti e cadherin, by Proteintech (13 mentions)
Anti vimentin, by Proteintech (12 mentions)
Anti n cadherin, by Proteintech (11 mentions)
Anti gapdh, by Proteintech (8 mentions)
Pvdf membrane, by Merck Group (7 mentions)
Anti β actin, by Proteintech (4 mentions)
Anti bcl 2, by Proteintech (4 mentions)
Anti gapdh, by Cell Signaling Technology (4 mentions)
Anti cdk6, by Proteintech (3 mentions)
Anti cyclin d1, by Proteintech (3 mentions)
Anti mmp7, by Proteintech (3 mentions)
Anti bax, by Proteintech (3 mentions)
Anti β catenin, by Proteintech (2 mentions)
Anti p akt, by Proteintech (2 mentions)
Anti akt, by Proteintech (2 mentions)
Anti cleaved caspase 3, by Cell Signaling Technology (2 mentions)
Anti vimentin, by Abcam (2 mentions)
Anti e cadherin, by Cell Signaling Technology (2 mentions)
Anti gapdh, by Abcam (2 mentions)
48 protocols using anti mmp9
1
Western Blot Analysis of EMT Markers
2
Western Blot Analysis of Protein Targets
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Whole-cell proteins were harvested, and the protein concentrations were measured using a BCA kit (Biosharp, Hefei, China). Equal amounts of protein (20 μg) were separated by 10% SDS-PAGE and transferred onto nitrocellulose membranes (EMD Millipore, MA, USA); subsequently, the membranes were blocked in 5% nonfat dry milk for 2 hours at room temperature. The membranes were incubated with primary antibodies at 4°C overnight, washed with TBST, and incubated with anti-HRP-conjugated Affinipure goat anti-rabbit/mouse lgG(H+L) (Proteintech, Wuhan, China). The signals were detected using an ECL Western blotting kit (Biosharp, Hefei, China). The primary antibodies used included anti-PADI6 (Thermo Fisher, MA, USA), anti-GAPDH (Proteintech, Wuhan, China), anti-YAP1 (Proteintech, Wuhan, China), anti-FLAG (Sigma, MO, USA), anti-beta tubulin (Proteintech, Wuhan, China), anti-14-3-3 (Proteintech, Wuhan, China), anti-MMP2 (Proteintech, Wuhan, China) and anti-MMP9 (Proteintech, Wuhan, China) antibodies.
3
Protein Expression Analysis by Western Blot
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Total cell extracts were subjected to 12% SDS-PAGE and separated proteins transferred to polyvinylidene fluoride membrane. Western blot was performed using standard protocols described previously [46 (link)]. Rabbit anti-DEK, anti-E-cadherin, anti-vimentin, anti-MMP2, and anti-MMP9 antibodies were purchased from Proteintech (Chicago, IL, USA). Mouse anti-GAPDH antibody was obtained from Chemicon (Temecula, CA, USA).
4
Protein Expression Analysis Protocol
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Protein extraction and blotting was performed as described previously [7 ]. Western blots were probed with the following antibodies anti-BMP5 (Abcam, ab88064, 1:500), anti-PCNA (Immunoway, 1:5000), anti-MMP2 (Proteintech, 1:500), anti-MMP9 (Proteintech, 1:1000), anti-E-cadherin (BD Biosciences, 1:1000), anti-EPSTI1 (Proteintech, 1:500) and anti-GAPDH (Immunoway, 1:5000).
5
Western Blot Analysis of EMT Markers
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Cells were lysed with RIPA lysate (Beyotime, Shanghai, China), and the concentration of protein was determined by the BCA assay kit (Solarbio, Beijing, China). Briefly, the equal amount of protein was separated with 10% SDS-PAGE (Epizyme, Shanghai, China), and transferred to PVDF membranes (Millipore, MA, USA). The PVDF membranes were blocked with 5% skim milk and incubated at 4 °C overnight with primary antibody, including anti-Fibronectin (Cell Signaling Technology, MA, USA), anti-N-cadherin (CST), anti-E-cadherin (CST), anti-MMP2 (CST), anti-MMP9 (CST), anti-Cytokeratin (Proteintech, Wuhan, China), anti-Vimentin (CST), anti-Ran (Proteintech), anti-Akt (CST), anti-p-Akt (CST), GSK3β (CST), anti-p-GSK3β (CST), anti-β-catenin (CST). β-actin (CST) was used as internal controls. Then the PVDF membranes were incubated with a secondary antibody for 1 h. The blots were scanned by Amersham Imager 600 (GE, Boston, MA, USA). Each sample was repeated three times and the three independent western blot bands were quantitatively analyzed by Image J software (NIH, Bethesda, Maryland, USA).
6
Protein Expression Analysis by Western Blot
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The cells under different conditions were harvested and protein was extracted. After quantification by BCA assay kit (KeyGEN, China), each protein sample (45 μg) was separated on a 10% SDS polyacrylamide gel, transferred to polyvinylidene difluoride polyscreen membrane (millipore, Germany) and then incubated with anti-MMP-9 (1 : 1000; Proteintech, China), anti-MMP-10 (1 : 500; Sigma-Aldrich, USA), anti-E-cadherin (1 : 500; Proteintech, China) anti-vimentin (1 : 300; Proteintech, China) and anti-β-actin (1 : 1000, Proteintech, China) antibodies overnight at 4 °C. Secondary anti-rabbit antibody (Proteintech, China) was added relative to the primary antibody and incubated for 2 h at room temperature. The blots were finally developed using an ECL western blot protocol (Lumilight enhanced chemiluminescence, Roche). Protein signals were visualized. Quantification of immune reactivity was performed by densitometric scanning using the Quantity One-1-Danalysis software (Bio-Rad, Hercules, CA).
7
Oxidative Stress Biomarker Assay Protocol
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Griess reagent, 5,5’-Dithiobis (2-nitrobenzoic acid) and Lucigenin were purchased from Sigma-Aldrich. Hydrogen peroxide 33% was procured from PanReac AppliChem. The following antibodies were used in this work: Anti-fibronectin was obtained from GeneTex. Anti-MMP-9 and anti-Bcl2 were obtained from Proteintech. Anti-vimentin, anti-MMP-3, Anti-Akt, Anti-p38, Anti-Cleaved PARP1, Anti-p53 (acetyl K382), Anti-JNK, Anti-alpha smooth muscle Actin, Anti-NF-kB p65 (phospho S536), and anti-β catenin were all obtained from Abcam (Cambridge, UK). Anti-cleaved caspase-3, anti-cleaved aspase-9, and anti-cleaved caspase-8 were obtained from Cell Signaling. Rabbit Polyclonal JNK1/2/3 was obtained from ORIGENE. Anti-beta Actin was obtained from Thermo Fisher Scientific. Peroxidase AffiniPure Goat Anti-Rabbit IgG and Peroxidase AffiniPure Goat Anti-Mouse IgG were obtained from Jackson ImmunoResearch Inc. The following ELISA kits were used in this work: nitrotyrosine ELISA kit (Abcam, ab113848), Hydrogen Peroxide Colorimetric/Fluorometric assay kit (Biovision, K265-200), malondialdehyde assay kits (Northwest Life Sciences Specialties, NWK-MDA01) and Superoxide Dismutase assay kit (Cayman chemical, 706002).
8
Antibody Profiling for Cell Signaling
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The antibodies were as follows: anti-Rab32 (Proteintech, #10999-1-AP, 1:1000), anti-MMP2 (Proteintech, #10373-2-AP, 1:1000), anti-MMP9 (Proteintech, #10375-2-AP, 1:1000), anti-β-Actin (CST, #4970, 1:1000), anti-N-cadherin (CST, #13116, 1:1000), anti-E-cadherin (CST, #3195, 1:800), anti-Vimentin (CST, #5741, 1:1000), anti-p-Drp1(Ser616) (CST, #3455, 1:1000), anti-p-Drp1(Ser637) (Abmart, #TD2980, 1:1000), anti-Drp1(CST, #8570, 1:1000), anti-Tom20 (Abcam, #ab56783, 1:1000), anti-p-ERK1/2 (CST, #4370, 1:2000), anti-ERK1/2 (CST, #4695, 1:2000). All secondary antibodies (HRP-conjugated anti-rabbit and anti-mouse, #A0208 and #A0216, 1:2000) were purchased from Beyotime Biotechnology (Shanghai, China). Anti-HA-tag (CST, #3724, 1:50 for IP and 1:1000 for western blot) and anti-IgG antibody (Servicebio, #GB23301, 1:100 for IP). Mdivi-1 (#S-7162), ERK1/2 inhibitor SCH772984 (#S-7101) were purchased from Selleck.
9
Immunofluorescence Analysis of Brain Markers
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The cultured cells or brain samples were fixed with 4% paraformaldehyde. Then, cells or coronal brain sections (30 μm) of the frontal cortex were incubated with the following primary antibodies at 4°C overnight: anti-MMP-9 (1:200; Proteintech, 10375-2), anti-MMP-9 (1:100; Santa Cruz Biotechnology, sc21733), anti-GFAP (1:1000; Abcam, ab4674), anti-NeuN (1:500; Abcam, ab104224), anti-myeloperoxidase (MPO; 1:100; Proteintech, 22225-1), anti-ionized calcium binding adapter 1 (IBA1; Synaptic Systems, 234004, 1:400), anti-CD31 (1:200; Novus Biologicals, NBP1-71663), anti-NDRG2 (1:200; Abcam, ab174850), anti-NDRG2 (1:2000; Proteintech, 67191-1), anti-p-Smad2(Ser465/467)/Smad3(Ser423/425) (1:200; Cell Signaling Technology, 8828), anti-PPM1A (1:200; Abcam, ab14824), and anti–laminin α2 (1:200; Sigma-Aldrich, L0663). Samples were then incubated with the secondary antibodies of Alexa Fluor 488/594 Donkey anti-Mouse/Rabbit/Chicken IgG (1:400; Invitrogen) at room temperature for 4 hours. 4′,6-Diamidino-2-phenylindole staining was then used to label cellular nuclei. Last, images were acquired by laser confocal microscopy (Nikon, A1, Tokyo, Japan). When red and green were double-labeled in images, red was replaced by purple pseudo-color.
10
Western Blot Analysis of Colon Cancer Proteins
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Protein was extracted from colon tissues or colon cancer cell lines with radioimmunoprecipitation assay (RIPA) buffer with a proteinase inhibitor. The Protein BCA Assay Kit (Bio-Rad) was used to measure the protein concentration in the lysate. 20 μg of protein were separated by SDS-PAGE (80 V, 2.5 h) and then transferred to polyvinylidene fluoride (PVDF) membranes (Millipore, USA). After that, the PVDF membranes were treated with a primary antibody (4°C, overnight). Followed by treatment with a peroxidase-conjugated secondary antibody, all the blots were visualized using ECL solutions and quantified. The following antibodies were used: anti-MMP-2 (1:500, Proteintech, Wuhan, Hubei, China), anti-MMP-9 (1:500, Proteintech), anti-CXCR-4 (1:2000, Proteintech), anti-vimentin (1:2000, Proteintech), anti-N-cadherin (1:2000, Proteintech), anti-E-cadherin (1:1000, Cell Signaling Technology), anti-DLCK3 (1:1000, Boster Bio, CA, USA) and anti-GAPDH (1:1000, Abcam, Cambridge, UK).
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