The harvested cell aggregate, native and decellularized tissue sections from porcine heart and TA muscle from rat were fixed in 4% phosphate-buffered paraformaldehyde for 24 hours, embedded in paraffin. Eight-micrometer-thick serial sections were cut from the paraffin-embedded blocks and underwent H&E staining and Masson’s Trichrome staining. Immunohistochemical staining was performed using standard procedures as described previously [35 (link), 36 (link)]. Briefly, sections were incubated with primary antibodies as follows: anti-Col-I (1:200; Santa Cruz, USA), anti-fibronectin (1:200; Abcam) and anti-integrin-β1 (1:200; Abcam). The same source IgG was used for the negative control instead of the primary antibodies. Biotinylated secondary antibodies (1:1000) were purchased from Sigma-Aldrich. The stained sections were observed using the light microscope (Nikon, Japan). The photographs were evaluated by Image-Pro Plus 6.0 (Media Cybernetics, USA) from three randomly selected views of each specimen.
Anti integrin β1
Manufactured by Abcam
Sourced in United Kingdom, United States
Anti-integrin β1 is a lab equipment product that recognizes the integrin β1 protein. Integrins are cell surface receptors that mediate cell-cell and cell-extracellular matrix interactions. This product can be used to study the expression and function of integrin β1 in various biological systems.
Automatically generated - may contain errors
Lab products found in correlation
Anti pten, by Abcam (3 mentions)
Anti akt, by Abcam (2 mentions)
Anti vinculin, by Abcam (2 mentions)
Anti p akt, by Abcam (2 mentions)
Hrp conjugated secondary antibody, by Abcam (2 mentions)
Fitc conjugated isotype control, by BD (2 mentions)
Fitc conjugated anti α2 integrin, by BioLegend (2 mentions)
Anti cd24, by Santa Cruz Biotechnology (2 mentions)
Biotinylated secondary antibody, by Merck Group (1 mentions)
Anti col 1, by Santa Cruz Biotechnology (1 mentions)
Anti fibronectin, by Abcam (1 mentions)
Image pro plus 6, by Media Cybernetics (1 mentions)
Anti integrin α5, by Abcam (1 mentions)
Anti integrin α5β1 antibody, by Merck Group (1 mentions)
Nitrocellulose filter membrane, by Bio-Rad (1 mentions)
Anti phospho akt, by Abcam (1 mentions)
Anti phospho pi3k, by Abcam (1 mentions)
Bca protein assay kit, by Beyotime (1 mentions)
Anti pi3k, by Abcam (1 mentions)
Alexa fluor 594 goat anti rabbit igg, by Thermo Fisher Scientific (1 mentions)
22 protocols using anti integrin β1
1
Histological Analysis of Tissue Samples
2
Integrin-Mediated Cell Adhesion Assay
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FN‐mediated cells in the first 12 h were pretreated with 10 µg mL−1 Anti‐Integrin α5 (Abcam), Anti‐Integrin β1 (Abcam), Anti‐Integrin α5β1 antibody (Millipore), or noninhibitory isotype control antibody (Millipore) for 30 min at 37 °C before washing in PBS. The cells (2 × 104 cells cm−2) were seeded in FN‐coated dishes for 1 h at 37 °C. After removal of nonadherent cells by washing with PBS, adherent cells were harvested with trypsin and quantified in triplicates with a hemocytometer. In addition, the area of adherent cells was quantified with the ImageJ software (NIH, version 1.25p).
3
Western Blot Analysis of Integrin β1, PI3K, and Akt
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Cells were lysed in cold RIPA lysis buffer supplemented with Protease Inhibitors cocktail (CST Inc., USA). The protein concentration was quantified using BCA protein assay kit (Beyotime, China) following the manufacturer’s introduction. Twenty micrograms of protein were mixed with 1× loading buffer and boiled for 5 mins at 100°C and were resolved by 8% or 10% SDS-PAGE and transferred to the nitrocellulose filter membrane (Bio-Rad). Target protein was detected using specific primary antibody, including: anti-Integrin β1 (1:1000; Abcam, Cambridge, UK), anti-PI3K (1:1000; Abcam), anti-Akt (1:1000; Abcam), anti-phospho-PI3K (1:1000; Abcam), anti-phospho-Akt (1:1000; Abcam). All experiments were repeated three independent times and the target protein level was quantified by ImageJ and normalized to internal control.
4
Immunofluorescence Analysis of Cell Markers
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Cells were fixed with 4% paraformaldehyde and permeabilised with Triton X-100 before blocking of nonspecific binding sites with 10% goat serum. Then, the cells were incubated overnight with rabbit monoclonal [SP6] anti-Ki67 (Abcam, Cambridge, UK) or anti-Vinculin (Abcam, Cambridge, UK) or anti- Integrin-β1 (Abcam, Cambridge, UK) at 4 °C, followed by incubation with Alexa Fluor 594 goat anti-rabbit IgG (1:200; Molecular Probes, Eugene, OR, USA) for 2 h at 37 °C. Cells were counterstained with DAPI to visualise nuclei and phalloidin to visualise actin. Images were captured using a Nikon confocal laser scanning microscope.
5
Immunohistochemical Analysis of NSCLC
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The NSCLC tissues were fixed in 4% paraformaldehyde for 72 h and then sectioned. The samples were deparaffinized and rehydrated using alcohol and water. For antigen recovery, the samples were treated with sodium citrate buffer at 100℃ for 5 min. For blocking, 5% BSA was used. Subsequently, the samples were incubated overnight at 4℃ with primary antibodies: anti-fibrin α subunit (1:100; Abcam, Cambridge, UK), anti-p-AKT (1:100; Abcam, Cambridge, UK), anti-integrin β1 (1:100; Abcam, Cambridge, UK), and anti-PTEN (1:200; Abcam, Cambridge, UK). Finally, the detection was carried out using the ABC HPR Kit (Thermo Fisher, MA, USA).
6
Probing PI3K/AKT Signaling in Bladder Cancer
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We fixed and permeabilized MB49 cells treated with collagen or E7820 to examine the PI3K/AKT signaling pathway in bladder cancer cells. Then the cells were labeled with anti‐integrin β1 (1:200; Abcam), anti‐p‐PI3K (1:200; Abcam), or p‐AKT (1:200; Abcam) followed by Alexa 594 secondary Abs (1:800; Abcam). In addition, nuclei were labeled with DAPI. All immunofluorescence images were captured from FV1000 laser scanning confocal microscope (Leica, Wetzlar, Germany).
7
Western Blot Analysis of Signaling Proteins
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For Western blot analysis, the following antibodies were used (working dilutions are given in brackets). Rabbit polyclonal antibodies: anti-P(S473)-AKT (1:2000, D9E, Cell Signaling), anti-Bcl-3 (1:1000, C-14, Santa Cruz), anti-P(Thr202, Tyr204)-ERK1/2 and anti-ERK1/2 (both 1:2000, Cell Signaling), anti-ERα (1:2000, Santa Cruz, HC-20), anti-IGF1Rβ (1:2000, Cell Signaling), anti-P(Tyr705)-STAT3 (1:1000, D3A7, Cell Signaling) and anti-STAT3 (1:1000, 79D7, Cell Signaling); rabbit monoclonal antibodies: anti-integrin β1 (1:2000, EPR1040Y, Abcam), anti-GAPDH (1:5000, Ambion) and Ki67 (1: 2000, Epitomics, clone EPR3610); mouse monoclonal antibodies: anti-(pan)AKT (1:1000, 40D4, Cell Signaling), anti-E-cadherin (1:5000, BD Transduction Lab.) and anti-HIF1α (1:1000, BD Transduction Lab.). Anti-CAIX was kindly provided by S. Pastorekova. Secondary antibody conjugates (anti-rabbit/anti-mouse horse radish peroxidase, 1:2000) were purchased from Cell Signaling.
Fulvestrant (LKT Laboratories) was purchased from Biomol (Hamburg/Germany), PQ404 from Calbiochem and recombinant human insulin was from Sigma-Aldrich.
Fulvestrant (LKT Laboratories) was purchased from Biomol (Hamburg/Germany), PQ404 from Calbiochem and recombinant human insulin was from Sigma-Aldrich.
8
Integrin β1 Signaling Pathway Analysis
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The total proteins were extracted by radioimmunoprecipitation assay (RIPA) lysis buffer (Beyotime, Shanghai, China) with the protease inhibitor phenylmethylsulfonyl fluoride (PMSF) (Beyotime, Shanghai, China). Cell lysates were separated on 10% sodium dodecyl sulfate-polyacrylamide gels (SDS-PAGE), then blocked and incubated with the corresponding primary antibodies: anti-integrin β1 (1:1000, Abcam, Cambridge, UK), anti-phosphorylated FAK (1:1000, Abcam, Cambridge, UK), anti-FAK (1:1000, Abcam, Cambridge, UK), anti- phosphorylated ERK1/2 (1:1000, Abcam, Cambridge, UK), anti-ERK1/2 (1:1000, Abcam, Cambridge, UK), anti-NF-κB (1:1000, Abcam, Cambridge, UK). Subsequently, samples were incubated with an HRP-conjugated secondary antibody (1:1000, Abcam, Cambridge, UK). β-actin served as an internal control.
9
Ursolic acid and aspirin combination protocol
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Ursolic acid (UA, purity > 90%) was purchased from Xi'an Ocean Biological Engineering Co. (Xian, China). Aspirin (Asp, purity > 90%) was purchased from Aladdin Reagent Company (Shanghai, China). UA, Asp and Asp-UA were dissolved in dimethyl sulfoxide (DMSO) and were used in all experiments. PE-labeled mouse anti-human E-cadherin, vimentin, EpCAM and integrin α1, α3, α5, α6 and β1 integrin mAbs, PE/FITC mouse IgG1 kappa isotype control and APC mouse IgG1 kappa isotype control antibody were all purchased from Becton Dickinson (BD) PharmingenTM (New Jersey, USA). Mouse anti-human beta-actin (β-actin) antibody was purchased from Cell Signaling Technology (Danfoss, USA). Matrigel were obtained from BD BiocoatTM (New Jersey, USA). The primary anti-EGFR, anti-ERK, anti-MMP-2, anti-MMP-9, anti-COX-2, anti-E-cadherin, anti-β-catenin, anti-integrin α6, anti-integrin β1, anti-PTEN and anti-CD44 antibodies were obtained from Abcam Biotechnology (Cambridge, UK). The secondary antibody goat-anti-mouse-IgG or goat-anti-rabbit-IgG horseradish peroxidase was obtained from Promega (Madison, USA). All other reagents used in this study were of the highest purity commercially available.
10
Integrin Signaling in Melanocytes
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Melanocyte monocultures and cocultures using inserts for seven days were reacted with anti-integrin β1 (10 ug/ml; Abcam), anti-bFGF, or anti-SCF antibody for another day. These cells were transferred to each well (3×104 cells in 400 µl DMEM medium without FBS) of a 24-well tissue culture plate coated with fibronectin or laminin (CORNING), incubated for 30 minutes, and then washed with sterile PBS (pH 7.4). After fixing in 4% paraformaldehyde and staining nuclei with Hoechst 33258, the number of stained cells was counted in a fixed area using a standard light microscope (DM LB microscope; Leica Microsystems, Wetzla, Germany).
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