Angiosarcoma cells were incubated with lysis buffer (Complete Lysis-M, Roche Applied Science, Indianapolis, IN, USA). The lysate protein concentration was measured using a BCA Protein Assay kit (Pierce, Rockford, IL, USA). Equal amounts of protein (20 µg) were dissolved in NuPage LDS Sample Buffer (Invitrogen) and 10% NuPage Sample Reducing Agent (Invitrogen). Lysates were boiled at 98°C for 2 minutes and loaded and run on 4-12% NuPage Bis-Tris Gels (Invitrogen) at 200 V for 40 minutes. The proteins were transferred onto polyvinylidene fluoride membranes (Invitrogen) and blocked in 2% bovine serum albumin in 0.1% Tween-20 (Sigma-Aldrich) and Tris-buffered saline. The membranes were probed with anti-FOXM1 antibody (1:100, Santa Cruz) overnight at 4°C. The secondary antibody used was anti-rabbit horseradish peroxidase-conjugated IgG antibody. Anti-human actin mouse monoclonal antibody (1:4000, Millipore) was used as a loading control. Protein bands were detected using the ImageQuant LAS 4000 system (GE Healthcare).
Anti human actin mouse monoclonal antibody
Manufactured by Merck Group
Sourced in United Kingdom
The anti-human actin mouse monoclonal antibody is a laboratory reagent used for the detection and quantification of actin, a structural protein found in eukaryotic cells. This antibody is specific to the human form of actin and can be used in various immunoassay techniques, such as Western blotting, immunohistochemistry, and ELISA, to identify and measure actin levels in biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Polyvinylidene fluoride membrane, by Thermo Fisher Scientific (1 mentions)
Anti foxm1 antibody, by Santa Cruz Biotechnology (1 mentions)
Nupage bis tris gel, by Thermo Fisher Scientific (1 mentions)
Nupage sample reducing agent, by Thermo Fisher Scientific (1 mentions)
Imagequant las 4000 system, by GE Healthcare (1 mentions)
Nupage lds sample buffer, by Thermo Fisher Scientific (1 mentions)
Complete lysis m, by Roche (1 mentions)
Tween 20, by Merck Group (1 mentions)
Bovine serum albumin (bsa), by Merck Group (1 mentions)
3 protocols using anti human actin mouse monoclonal antibody
1
FOXM1 Protein Expression in Angiosarcoma
2
Western Blot Analysis of FOXM1
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The cells were washed twice with ice-cold phosphate-buffered saline (PBS), scraped, and collected in a microcentrifuge tube. Whole cell lysates were prepared from the cell lines. Anti-FOXM1 (1:200 dilution) antibody (R&D Systems). Anti-human actin mouse monoclonal antibody (1:5000; Millipore) was used as a loading control. The subsequent Western blot procedure was performed as described [33 (link)].
3
FOXM1 Protein Expression Analysis
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Whole cell lysates were prepared from transfected cell lines. To confirm the down-regulation of FOXM1, we evaluated the expression of FOXM1 protein by Western blot assay as described previously [37] . A total of 20 µg protein from each sample was used, and incubated with anti-FOXM1 (1:200 dilution) antibody. Anti-human actin mouse monoclonal antibody (1:5000; Millipore, Bedford, MA) was used as an internal control. Protein levels were standardized by actin, which was assigned an arbitrary level of 10, and the expression signal relative to this was taken as the expression value for each sample.
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