Cell seeding, collection of protein and western blot methods were previously described (22 (link), 23 (link)). Membranes were probed with the following antibodies: anti-MT1-MMP (Millipore, MAB3328), anti-GAPDH (Santa Cruz Biotechnology, C65), anti-FAK-(pY397) (BD Biosciences, 611722), anti-FAK (Cell Signaling, D2R2E), anti-ITGB1 (Cell Signaling, D2E5), anti-Cleaved-PARP (Cell Signaling, D64E10), anti-SMAD3-(pS423/S425) and anti-SMAD3 (EP568Y) (Abcam).
Anti itgb1
Manufactured by Cell Signaling Technology
Sourced in United Kingdom
Anti-ITGB1 is a laboratory reagent produced by Cell Signaling Technology. It is a monoclonal antibody that recognizes the ITGB1 (Integrin Beta 1) protein. ITGB1 is a subunit of various integrin receptors and plays a role in cell-cell and cell-extracellular matrix interactions.
Automatically generated - may contain errors
Lab products found in correlation
Anti itgb3, by Cell Signaling Technology (3 mentions)
Anti itgav, by Cell Signaling Technology (2 mentions)
Anti cd47, by Abcam (2 mentions)
Anti β actin, by Merck Group (2 mentions)
Anti cleaved parp, by Cell Signaling Technology (1 mentions)
Anti gapdh, by Santa Cruz Biotechnology (1 mentions)
Anti mt1 mmp, by Merck Group (1 mentions)
Anti fak, by Cell Signaling Technology (1 mentions)
Anti fak py397, by BD (1 mentions)
Polyvinylidene difluoride membrane, by Merck Group (1 mentions)
Anti flag, by Merck Group (1 mentions)
Anti shp2, by Cell Signaling Technology (1 mentions)
Anti phospho shp2, by Cell Signaling Technology (1 mentions)
Anti shp 1, by Cell Signaling Technology (1 mentions)
Anti phospho shp1, by Cell Signaling Technology (1 mentions)
Camk2, by Cell Signaling Technology (1 mentions)
Anti angptl2, by R&D Systems (1 mentions)
Anti phospho creb, by Cell Signaling Technology (1 mentions)
8 well chamber slide, by EQT (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Vector Laboratories (1 mentions)
7 protocols using anti itgb1
1
Protein expression and signaling analysis
2
Western Blot Analysis of Signaling Proteins
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Whole cell lysates were electrophoresed on 10% sodium dodecyl sulfate polyacrylamide gels and transferred onto polyvinylidene difluoride membranes (Millipore). The membranes were incubated with primary antibodies overnight at 4°C followed by incubation with appropriate horseradish peroxidase-conjugated secondary antibodies. The following antibodies were used: anti-CREB, anti-Phospho-CREB (Ser133), anti-SHP-1, anti-Phospho-SHP1 (Tyr564), anti-SHP-2, anti-Phospho-SHP2 (Tyr580), anti-ITGB1, anti-ITGB3, p-CaMK2, CaMK2 (Cell Signaling Technology), anti-ANGPTL2 (R&D Systems), anti-CaMK1 (Abcam), anti-p-CaMK4 (Abmart), anti-CaMK4 (Genscript), anti-Flag (Sigma), anti-GAPDH, and anti-β-actin (Calbiochem). Anti-LILRB2-PE (eBioscience) and isotype control of IgG-PE were used to detect the expression of LILRB2 on the different lung cancer cell lines and analyzed by flow cytometry.
3
Immunofluorescence Analysis of Cell Adhesion Molecules
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Cells (1 × 104/well) were seeded on 8-well chamber slides (Lab-Tech) after transfection. The cells were fixed, permeated and blocked. Then, they were incubated with the following antibodies: anti-VIM (1:200, Abcam), anti-ITGAV (1:1000, Cell Signaling Technology), anti-ITGB1 (1:1000, Cell Signaling Technology), anti-ITGB3 (1:500, Cell Signaling Technology) or anti-CD47 (1:200, Abcam). The secondary antibody, Alexa Fluor 488-conjugated goat anti-mouse IgG or anti-rabbit IgG (1:500, Invitrogen), was applied for 1 hour. Slides were mounted with antifading solution containing 4′-6′-diamidino-2-phenyl-indole (Vector Laboratories). Images were taken using a confocal microscope (Zeiss). All experiments were conducted in triplicate.
4
Western Blot Analysis of Integrin and Signaling Proteins
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Protein samples were separated by electrophoresis in SDS-polyacrylamide gel (6%) and transferred to nitrocellulose membrane. The membrane was blocked with a TBST buffer (1X TBS, 0.25% Tween-20) containing 5% skimmed milk at room temperature for 60 minutes under stirring. The primary antibodies used were: anti-ITGA5 (Cell Signaling, #98204) at dilution 1:1000, anti-ITGB1 (Cell Signaling, #4706) at dilution 1:1000, anti-tubulin (ABCAM, ab7291) at dilution 1:10000, anti-phospho-AKT (Cell Signaling, #4060) at dilution 1:2000, anti-AKT (Cell Signaling, #4691) at dilution 1:1000 and anti-GAPDH (Cell Signaling, #5174) at dilution 1:1000. Images were captured by ChemiDoc™ Imaging System from Bio-Rad.
5
Western Blotting Analysis of Protein Expression
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Total protein was extracted from tissues and transfected cells using RIPA buffer including protease inhibitor cocktail (Cell Signaling Technology), and western blotting was carried out as described previously [42 (link)]. The primary antibodies used were as follows: anti-DHFR (1:400, Cell Signaling Technology), anti-ZEB2 (1:1000, Abcam), anti-E-cadherin (1:250, Abcam), anti-VIM (1:1500, Cell Signaling Technology), anti-ITGAV (1:1000, Cell Signaling Technology), anti-ITGB1 (1:2000, Cell Signaling Technology), anti-ITGB3 (1:2000, Cell Signaling Technology), anti-CD47 (1:2000, Abcam), and anti-β-actin (1:5000, Sigma-Aldrich).
6
Western Blot Analysis of Osteoclast Markers
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The cells were first washed three times with PBS and lysed with radioimmunoprecipitation assay buffer with complete protease inhibitor (Solarbio). After protein quantification, the samples were separated by 12% SDS-PAGE and transferred to polyvinylidene difluoride membranes (Bio-Rad) according to the standard protocol. The membranes were blocked with 5% skim milk and incubated overnight at 4 °C with the appropriate primary antibodies. The primary antibodies were as follows: anti-TRAP, anti-CTSK, anti-NFATc1, and anti-Itgb1 (1:1000; Cell Signaling Technology). β-Actin was obtained from Affinity Biosciences. The membranes were subsequently incubated with secondary antibodies for 1 h at RT and washed with a Tris-buffered saline with Tween-20 for 30 min. The blots were visualized using Tanon image scanning. ImageJ software was used for quantitative analysis of the blots.
7
Protein Extraction and Western Blot Analysis
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Protein extraction and Western blot were carried out as described previously [12 (link)]. Briefly, for secretory protein collection, media supernatant was treated with the protease inhibitor cocktail. Anti-β-Actin (1:3000, A5441-.2 ML, Sigma-Aldrich, MO, USA), anti-QSOX2 (1:1000, ab121376, Abcam, Cambridge, UK), anti-ITGB1 (1:1000, #4706, Cell Signaling, MA, USA), anti-pAKT T308 (1:1000, #9275S, Cell Signaling, MA, USA), anti-pAKT S473 (1:1000, #9271S, Cell Signaling, MA, USA), and anti-total AKT1 (1:1000, #9272S, Cell Signaling, MA, USA) were used as primary antibodies. Goat anti-rabbit (A120-101P, Bethyl, TX, USA) and Goat anti-Mouse (Bethyl, A90-116P, TX, USA) were used as secondary antibodies.
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