Proteins were quantified by immunoblot analysis as described previously (21 (link), 24 (link)). For co-culture experiments, leukemia cells were collected by combining cells in the supernatant and after two washes with PBS. Antibodies against p-CRKL, BID, and BCL-XL were purchased from Cell Signaling Technology, MCL-1 from Santa Cruz Biotechnology Inc., BIM and CRKL from Abcam, BAX from Sigma, and BCL-2 from Dako. Signals were detected using an Odyssey infrared imaging system and quantitated using the Odyssey software program (version 3.0; LI-COR Biotechnology). β-ACTIN (Sigma) was used as a loading control.
Odyssey software program
Manufactured by LI COR
Sourced in United States
The Odyssey software program is a data analysis tool designed for use with LI-COR's imaging systems. It provides a suite of tools for processing and managing data collected from these systems. The software enables users to visualize, quantify, and analyze images generated by LI-COR's imaging hardware.
Automatically generated - may contain errors
Lab products found in correlation
Odyssey infrared imaging system, by LI COR (2 mentions)
Mcl 1, by Santa Cruz Biotechnology (1 mentions)
Bcl 2, by Agilent Technologies (1 mentions)
P crkl, by Cell Signaling Technology (1 mentions)
Bcl2 associated x (bax), by Merck Group (1 mentions)
Bcl xl, by Cell Signaling Technology (1 mentions)
β actin, by Merck Group (1 mentions)
Ab9484, by Abcam (1 mentions)
Ab131607, by Abcam (1 mentions)
Ab11304, by Abcam (1 mentions)
Irdye goat anti mouse, by LI COR (1 mentions)
Tgx precast gel, by Bio-Rad (1 mentions)
Sc 59886, by Santa Cruz Biotechnology (1 mentions)
Sc 507, by Santa Cruz Biotechnology (1 mentions)
Ab3242, by Merck Group (1 mentions)
4 protocols using odyssey software program
1
Quantifying Apoptosis-related Proteins
2
Western Blot Protein Quantification
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Protein levels were determined by western blot analysis as previously described.37 (link) Antibodies against β-catenin (#8480), CD44 (#5640) and p-CRKL (#3181) were purchased from Cell Signaling Technology (Danvers, MA, USA), and survivin (AF886) from R&D Systems (Minneapolis, MN, USA). β-Actin was used as a loading control. Proteins were visualized with Odyssey infrared imaging system and quantitated using the Odyssey software program (version 3.0; LI-COR Biotechnology, Lincoln, NE, USA).
3
Quantifying Protein Expression Levels
4
Western Blot Analysis of Protein Targets
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The Western blot procedure was consistent with our previous work and detailed elsewhere (Hüttemann et al., 2012 (link), 2013 (link); Lee et al., 2012 (link); Malek et al., 2013 (link)). Samples were loaded onto 7.5% (TSP-1, PGC-1α, PGC-1β, and ADAMTS-1) or 12% TGX pre-cast gels (Bio-Rad, Hercules, CA, USA).
The mouse monoclonal primary antibodies used were TSP-1 (1:500, sc-59886, Santa Cruz Biotechnology, Inc), CD47 (1:500; 3847-1, Epitomics), PGC-1β (1:100; sc-373771, Santa Cruz Biotechnology, Inc), α-tubulin (1:2,000, ab11304, Abcam), ADAMTS1 (1:500; sc-47726, Santa Cruz Biotechnology, Inc), and GAPDH (1:2,000, ab9484, Abcam). The polyclonal primary antibodies used were VEGF (1:500, sc-507, Santa Cruz Biotechnology, Inc), VEGFR2 (1:500; 2479, Cell Signaling), FoxO1 (1:200; 2880, Santa Cruz Biotechnology, Inc), Anti-TFAM (1:1,000; ab131607, Abcam), and PGC-1α (1:1,000; AB3242, Millipore). The secondary antibodies used were goat anti-mouse IRDye (1:30,000) and goat anti-rabbit IRDye (1:30,000) purchased from Li-Cor Biosciences. Loading control for target proteins were normalized to α-tubulin or GAPDH. Quantification of bands were analyzed with the Odyssey software program (Li-Cor Biosciences).
The mouse monoclonal primary antibodies used were TSP-1 (1:500, sc-59886, Santa Cruz Biotechnology, Inc), CD47 (1:500; 3847-1, Epitomics), PGC-1β (1:100; sc-373771, Santa Cruz Biotechnology, Inc), α-tubulin (1:2,000, ab11304, Abcam), ADAMTS1 (1:500; sc-47726, Santa Cruz Biotechnology, Inc), and GAPDH (1:2,000, ab9484, Abcam). The polyclonal primary antibodies used were VEGF (1:500, sc-507, Santa Cruz Biotechnology, Inc), VEGFR2 (1:500; 2479, Cell Signaling), FoxO1 (1:200; 2880, Santa Cruz Biotechnology, Inc), Anti-TFAM (1:1,000; ab131607, Abcam), and PGC-1α (1:1,000; AB3242, Millipore). The secondary antibodies used were goat anti-mouse IRDye (1:30,000) and goat anti-rabbit IRDye (1:30,000) purchased from Li-Cor Biosciences. Loading control for target proteins were normalized to α-tubulin or GAPDH. Quantification of bands were analyzed with the Odyssey software program (Li-Cor Biosciences).
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