To further characterize the primary cells, total proteins were obtained from the adherent cells at passage 3 by adding ice‐cold RIPA (radioimmunoprecipitation assay) lysis buffer (Servicebio). The equivalent protein (25 μg) was loaded to an SDS‐PAGE gel and then imprinted onto a polyvinylidene fluoride membrane. The membrane was blocked with 5% skim milk for 1.5 h at room temperature and incubated with a specific primary antibody and an appropriate secondary antibody at −4°C overnight. Antibodies used included the following: anti‐CD31 (bs‐0195R, Bioss Antibodies, 1:500), anti‐vimentin (V6389, Sigma‐Aldrich, 1:2000), anti‐α‐SMA (A2547, Sigma‐Aldrich, 1:1500), anti‐collagen XV (bs‐0547R, Bioss Antibodies, 1:1500) and anti‐β‐actin (bs‐0061r, Bioss Antibodies, 1:5000). The bands on the membrane were developed via a Super ECL (enhanced chemiluminescence) Plus kit (Boster Biological Technology) and recorded via Image Lab software (4600SF, Tanon Science & Technology).
Anti vimentin v6389
Manufactured by Merck Group
Sourced in Sao Tome and Principe
Anti-vimentin (V6389) is a laboratory reagent used for the detection and analysis of the intermediate filament protein vimentin in biological samples. It is a mouse monoclonal antibody that specifically binds to vimentin, a cytoskeletal protein found in various cell types. The core function of this product is to serve as a tool for researchers and scientists to study the expression and distribution of vimentin in their research applications.
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Lab products found in correlation
2 protocols using anti vimentin v6389
1
Protein Expression Analysis of Primary Cells
2
Cytoskeletal protein analysis by Western blot
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The cytoskeletal isolation fractions were separated using 4% to 15% precast polyacrylamide gels (4568084; Bio-Rad, Hercules, CA, United States). The gels were then transferred onto nitrocellulose membranes (1704271; Bio-Rad, Hercules, CA, United States) using a transfer system (Trans-Blot Turbo; 1705150; Bio-Rad, Hercules, CA, United States). The blocking and developing of the membranes were carried out as previously described [44 (link)]. The antibodies used were as follows: anti-vimentin (V6389; Sigma Aldrich, St. Louis, MO, United States), pan-cytokeratin (ab8068; Abcam, Cambridge, UK), anti-β-actin (A1978; Sigma Aldrich, St. Louis, MO, United States) and Horseradish-peroxide-conjugated goat anti-mouse secondary antibodies (GtxMu-004-DHRPX; ImmunoReagents, Raleigh, NC, United States). Membranes were stripped after imaging vimentin and pan-keratin, and exposed for 2 min to confirm that the membranes had been stripped, before incubating with β-actin antibody as a loading control.
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