Anti-α-actinin-4 was from Santa Cruz Biotechnology, Inc (Dallas, TX, USA, Cat #: SC-49333); anti-CD11b was from CedarLane Laboratories Limited (Hornby, Ontario, Canada, Cat #: CL8941AP); anti-Fibronectin was from Sigma (St. Louis, MO, USA, Cat #: F3648); anti-Integrin α8 was from R&D Systems (Minneapolis, MN, USA, Cat #: AF4076); anti-Laminin α1 was a gift from Dr. Dale Abrahamson (KU Medical Center, Kansas City, KS, rat monoclonal 8B3); anti-Laminin α2 and anti-β actin were from Sigma (St. Louis, MO, USA, Cat #: L0663); anti-Laminin α5 was a gift from Dr. Jeff Miner (Washington University, St. Louis, MO); anti-p-FAK397 was from Assay Biotechnology (Sunnyvale, CA, USA, Cat #: A0925) and from Invitrogen (Carlslab, CA); anti-Total FAK was from Cell Signaling Technology (Danvers, MA, USA, Cat #: 3285). Anti-MMP-10 antibodies were from Millipore (Billerica, MA, USA, Cat # ABT 289). All Alexa-fluor conjugated secondary antibodies were from Invitrogen (Carlsbad, CA), including donkey anti-rat 488, donkey anti-rabbit 555, goat anti-rat 488, goat anti-rabbit 555, donkey anti-rabbit 488, and donkey anti-goat 568. The small molecular inhibitor for FAK activation, TAE226 was from Chem Scene (Monmouth Junction, NJ, Cat #CS-0594); the peptide inhibitor for NF-kappaB (SN-50) was from Calbiochem (now EMD Millipore, Billerica, MA, Cat #481480)
Anti laminin α2
Manufactured by Merck Group
Sourced in United States
Anti–laminin α2 is a laboratory reagent used to detect the presence of laminin α2 protein in biological samples. Laminin α2 is a structural component of the extracellular matrix and is important for cell adhesion and signaling. This reagent can be used in various analytical techniques, such as immunohistochemistry and Western blotting, to identify and localize laminin α2 in tissues or cell lysates.
Automatically generated - may contain errors
Lab products found in correlation
Anti α actinin 4, by Santa Cruz Biotechnology (2 mentions)
Anti integrin α8, by R&D Systems (2 mentions)
Anti mmp 9, by Santa Cruz Biotechnology (2 mentions)
M o m kit, by Vector Laboratories (2 mentions)
Triton x 100, by Merck Group (2 mentions)
Power block universal blocking reagent, by BioGenex (2 mentions)
Donkey anti rabbit 488, by Thermo Fisher Scientific (1 mentions)
Donkey anti goat 568, by Thermo Fisher Scientific (1 mentions)
Goat anti rat 488, by Thermo Fisher Scientific (1 mentions)
Anti total fak, by Cell Signaling Technology (1 mentions)
Donkey anti rat 488, by Thermo Fisher Scientific (1 mentions)
Donkey anti rabbit 555, by Thermo Fisher Scientific (1 mentions)
Tae226, by ChemScene (1 mentions)
Anti fibronectin, by Merck Group (1 mentions)
Anti β actin, by Merck Group (1 mentions)
Anti cd11b, by Cedarlane (1 mentions)
Anti cd31, by Thermo Fisher Scientific (1 mentions)
Ab4674, by Abcam (1 mentions)
Anti mmp9, by Proteintech (1 mentions)
Anti pax7, by Developmental Studies Hybridoma Bank (1 mentions)
7 protocols using anti laminin α2
1
Immunofluorescence Staining of ECM Proteins
2
Immunohistochemical Staining for Cell Markers
3
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.
4
Immunofluorescence Staining of Satellite Cells
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Cryosections described above were used for immunohistochemistry. For immunocytochemistry, primary satellite cells were cultured on eight‐well chamber slides (MATSUNAMI) coated with Matrigel (BD Biosciences, San Jose, California, www.bd.com ). Tissue sections or cells were fixed in 4% paraformaldehyde in PBS for 10 minutes at room temperature, and then permeabilized with 0.2% Triton X‐100 (Sigma‐Aldrich, St. Louis, Missouri, www.sigmaaldrich.com ) in phosphate buffered saline (PBS) for 15 minutes at room temperature. After blocking with Power Block Universal Blocking Reagent (BioGenex, Fremont, California, http://biogenex.comLaboratories ) or M.O.M. kit (Vector Laboratories, Burlingame, California, www.vectorlabs.com ), the fixed cells were incubated with primary antibodies overnight at 4°C. After washing, bound primary antibodies were labeled with fluorescence‐conjugated secondary antibodies for 1 hour at room temperature. The immunostained samples were mounted with Mounting medium for fluorescence with DAPI (Vector Laboratories). Primary and secondary antibodies were as follows: anti‐laminin α2 (Sigma‐Aldrich), anti‐Pax7 (Developmental Studies Hybridoma Bank, Iowa City, Iowa, http://dshb.biology.uiowa.edu ), anti‐Ki67 (Leica or BD Biosciences), anti‐MHC (Leica), and mouse/rabbit/rat IgG‐Alexa488, ‐Alexa594, or Alexa647 (Life Technologies, St. Aubin, France, www.lifetech.com ).
5
Western Blot Analysis of Brain Tissue
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The cell samples or frontal lobes of brain tissues were lysed with radioimmunoprecipitation assay buffer (Thermo Scientific, 87788), and 20 μg of samples was separated with 4 to 20% SDS–polyacrylamide gel electrophoresis and transferred to a polyvinylidene difluoride membrane (Millipore, Shanghai, China). After blocking, the membrane was incubated overnight with the indicated primary antibody at 4°C. The following primary antibodies were used: anti-GFAP (1:10,000; Abcam, ab7260), anti-NDRG2 (1:2000; Abcam, ab174850), anti-p-Smad2 (Ser465/467)/Smad3 (Ser423/425) (1:1000; Cell Signaling Technology, 8828), anti-Smad2/3 (1:1000; Cell Signaling Technology, 8685), anti-Smad3 (1:1000; Cell Signaling Technology, 9528), anti–MMP-9 (1:1000; Santa Cruz Biotechnology, sc21733), anti–laminin α2 (1:200; Sigma-Aldrich, L0663), anti-occludin (1:10000; Proteintech, 66378-1), anti–ZO-1 (1:1000; Proteintech, 66452-1), anti-flag (1:5000; Sigma-Aldrich, F1804), anti-hemagglutinin (HA; 1:20000; Proteintech, 66006-2), and anti–β-actin (1:5000; Abcam, ab8226). Horseradish peroxidase–conjugated secondary antibodies were used. Chemical reactions were detected with an ECL system (Advansta, Menlo Park, CA, USA). The scanned images were analyzed with ImageJ NIH software.
6
Immunohistochemistry and Immunocytochemistry Analysis
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Immunohistochemistry was performed using the cryosections described above. For immunocytochemistry, primary satellite cells were cultured on 8-well chamber slides (MATSUNAMI) coated with Matrigel. Tissue sections or cells were fixed in phosphate-buffered saline (PBS) containing 4% paraformaldehyde for 10 min at room temperature, and then permeabilized with PBS containing 0.2% Triton X-100 (Sigma–Aldrich) for 15 min at room temperature. After blocking with Power Block Universal Blocking Reagent (BioGenex) or a M.O.M. kit (Vector Laboratories), the fixed cells were incubated with primary antibodies overnight at 4 °C. After washing, bound primary antibodies were labeled with fluorescence-conjugated secondary antibodies for 1 h at room temperature. The immunostained samples were mounted with Mounting medium containing DAPI (Vector Laboratories). The primary and secondary antibodies were as follows: anti-laminin α2 (Sigma–Aldrich), anti-Pax7 (SantaCruz), anti-collagen I (Abcam), anti-collagen IV (Abcam), anti-collagen VI (Abcam) and mouse/rabbit/rat IgG-Alexa488 or -Alexa594 (Life Technologies).
7
Laminin Isoforms and Tight Junction Proteins in Brain
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Cortex and striatum were carefully dissected and immediately homogenized on ice. Total protein concentration was determined using the BCA Protein Assay Kit (Pierce 23,227). Equal amount of protein was loaded and separated in SDS-PAGE and transferred to PVDF membranes (Millipore). Next, the membranes were probed with primary antibodies [anti-Laminin-α2 (1:500, Sigma L0663), anti-Laminin α5 (1:800, generated as described in [47 (link)]), Claudin-5 (1:500, ThermoFisher 35–2500), ZO-1 (1:500, ThermoFisher 61–7300), and anti-GAPDH (1:1000, Abcam AB9484)] over night at 4 °C, followed by appropriate horseradish peroxidase-conjugated secondary antibodies [donkey anti-mouse (1:2500, Jackson ImmunoResearch Laboratory 715–035-151), donkey anti-rabbit (1:2500, Jackson ImmunoResearch Laboratory 711–035-152), and donkey anti-rat (1:2500, Jackson ImmunoResearch Laboratory 712–035-153] at room temperature for 1 h. Then, target proteins were visualized using the ChemiDoc Imaging System (Bio-Rad). For quantification, the density of target blots was normalized to that of GAPDH, and the expression of target proteins in α5-PKO brains was normalized to that in control brains. Four animals were used for quantification.
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