For osteogenesis relative protein, antibody BMP2 (1:1000; abcam, ab284387), RUNX2 (1:1000; abcam, ab236639), ALP (1:1000; abcam, ab203106) and OPN (1:1000; abcam, ab63856) were chosen. Osteogenesis signal pathway test was used PI3K/Akt (1:1000; abcam, ab283852) and p38/MAPK14 (1:1000; abcam, ab170099) antibody. Protein extraction and Western blot were carried out as previously described. The membrane was then washed and blocked with 5% skimmed milk and probed overnight. The bound primary antibody was detected by peroxidase-conjugated goat anti-mouse IgG (Sigma-Aldrich) and the ECL Western blot analysis system (Millipore). Each experiment was performed at least three times. Statistical significance was set at 5%.
Ecl western blot analysis system
Manufactured by Merck Group
Sourced in United States
The ECL Western blot analysis system is a laboratory equipment used for the detection and quantification of proteins in biological samples. It utilizes a chemiluminescent detection method to visualize and analyze the results of Western blot experiments.
Automatically generated - may contain errors
Lab products found in correlation
Ab236639, by Abcam (1 mentions)
Ab284387, by Abcam (1 mentions)
Ab63856, by Abcam (1 mentions)
Ab283852, by Abcam (1 mentions)
Peroxidase conjugated goat anti mouse igg, by Merck Group (1 mentions)
Ab170099, by Abcam (1 mentions)
Peroxidase conjugated goat anti rabbit immunoglobulin g, by Merck Group (1 mentions)
Anti cd81, by Santa Cruz Biotechnology (1 mentions)
Anti cd63, by Santa Cruz Biotechnology (1 mentions)
Myd88, by Cell Signaling Technology (1 mentions)
Goat anti rabbit igg, by Merck Group (1 mentions)
β actin, by Cell Signaling Technology (1 mentions)
Gapdh, by MultiSciences Biotech (1 mentions)
Traf6, by Abcam (1 mentions)
3 protocols using ecl western blot analysis system
1
Osteogenesis Protein Expression Analysis
2
Characterization of Nanoparticle Size and Exosome Markers
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The size distribution and concentration of the nanoparticles before and after treatment were characterized by NTA. Each NTA video was 60 s, and each result was averaged by at least three tests. The concentration of the preprocessed polydisperse sample in the NTA test was diluted to guarantee that there were 70 to 120 particles per frame to ensure statistics and suppress the overlapping effect. For monodisperse samples, at least 10 particles per frame were guaranteed.
For protein extraction and Western blot, plasma samples were lysed with lysis buffer and subjected to 10% SDS–polyacrylamide gel electrophoresis gels and transferred to a polyvinylidene difluoride membrane. Anti-CD63 (1:200; Santa Cruz Biotechnology) and anti-CD81 (1:200; Santa Cruz Biotechnology) antibodies were used as primary antibodies to confirm the presence of exosomes. The bound primary antibody was detected by peroxidase-conjugated goat anti-rabbit immunoglobulin G (Sigma-Aldrich) and the ECL Western blot analysis system (Millipore). Each experiment was performed at least three times.
For protein extraction and Western blot, plasma samples were lysed with lysis buffer and subjected to 10% SDS–polyacrylamide gel electrophoresis gels and transferred to a polyvinylidene difluoride membrane. Anti-CD63 (1:200; Santa Cruz Biotechnology) and anti-CD81 (1:200; Santa Cruz Biotechnology) antibodies were used as primary antibodies to confirm the presence of exosomes. The bound primary antibody was detected by peroxidase-conjugated goat anti-rabbit immunoglobulin G (Sigma-Aldrich) and the ECL Western blot analysis system (Millipore). Each experiment was performed at least three times.
3
Protein Extraction and Western Blot Analysis
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Protein extraction and western blot were carried out as previously described18 (link). Various amounts of protein prepared from cell lysates and TEXs were loaded onto SDS-polyacrylamide gel (10%). The membrane was then washed and blocked with 5% skimmed milk and probed for 1 h with different primary antibodies including mouse monoclonal antibodies: Alix (1:200; Cell signaling technology, USA) and GAPDH (1:1000; MultiSciences Biotech Co., Ltd, China); and rabbit polyclonal antibodies: β-actin (1:2000; Cell signaling technology, USA), CD63 (1:200, Santa Cruz, USA), HMGN1 (1:1000; ProteinTech, USA), AFP (1:2000; Abcam, UK), TLR4 (1:1000; ProteinTech, USA), MyD88 (1:1000; Cell signaling, USA), and TRAF6 (1:5000; Abcam, UK), respectively. The bound primary antibodies were detected by peroxidase-conjugated goat anti-mouse, rabbit anti-mouse, or goat anti-rabbit IgGs (1:5000; Sigma, USA), respectively, and the ECL western blot analysis system (Millipore, Billerica, MA) was used. Uncropped images of all western blots can be found in Supplementary Fig. 10 .
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