All other fine chemicals were purchased from either Sigma-Aldrich (St. Louis, MO, USA) or CalBiochem (Carlsbad, CA, USA). Bicinchoninic acid (BCA) reagents, horseradish peroxidase (HRP)-conjugated secondary antibody, Superblock (TBS), and enzyme-linked immunosorbent assay (ELISA) MaxiSorp 96-well plates were purchased from Thermo Fisher Scientific (Bedford, MA, USA). Amplex Red soluble fluorophore was purchased from Life Technologies (Carlsbad, CA, USA). The Bio-Plex Pro™ Human Cytokine 27-plex Assay was purchased from Bio-Rad (Hercules, CA, USA). The Luminex MAGPIX system was purchased from Luminex Corp. (Austin, TX, USA). The SpectraMax M5 microplate reader was purchased from Molecular Devices Corp. (Sunnyvale, CA, USA).
Amplex red soluble fluorophore
Manufactured by Thermo Fisher Scientific
Sourced in United States
Amplex Red soluble fluorophore is a highly sensitive and stable fluorescent probe used for the detection and quantification of hydrogen peroxide (H2O2) and other peroxides in biological samples. It is a non-fluorescent compound that becomes highly fluorescent upon oxidation, allowing for the sensitive measurement of peroxide levels.
Automatically generated - may contain errors
Lab products found in correlation
Spectramax m5 microplate reader, by Molecular Devices (7 mentions)
Elisa maxisorp 96 well plates, by Thermo Fisher Scientific (6 mentions)
Horseradish peroxidase hrp conjugated secondary antibody, by Thermo Fisher Scientific (5 mentions)
Rplpo antibody, by Proteintech (5 mentions)
Akt pathway total and phospho panels, by Thermo Fisher Scientific (2 mentions)
Hrp conjugated secondary antibody, by Thermo Fisher Scientific (1 mentions)
Bicinchoninic acid (bca), by Thermo Fisher Scientific (1 mentions)
Horseradish peroxidase, by Thermo Fisher Scientific (1 mentions)
Superblock, by Thermo Fisher Scientific (1 mentions)
Bio plex pro human cytokine 27 plex assay, by Bio-Rad (1 mentions)
Streptavidin alkaline phosphatase, by Vector Laboratories (1 mentions)
Maxisorp 96 well plate, by Thermo Fisher Scientific (1 mentions)
Choline chloride, by Thermo Fisher Scientific (1 mentions)
Daidzein, by Merck Group (1 mentions)
Genistein, by Merck Group (1 mentions)
7 protocols using amplex red soluble fluorophore
1
Comprehensive Reagents and Instruments
2
Quantification of Choline, Daidzein, and Genistein
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Choline chloride was purchased from Thermo Fisher (Waltham, MA, USA). Daidzein (>98% purity) and Genistein (>95% purity) were purchased from Sigma-Aldrich Company (St. Louis, MO, USA). Commercial antibodies, including their sources, concentrations or dilutions used, vendors, and RRID numbers, are listed in Table 5 . Bicinchoninic acid (BCA) reagents, horseradish peroxidase (HRP)-conjugated secondary antibody, superblock (TBS), and enzyme-linked immunosorbent assay (ELISA). MaxiSorp 96-well plates were purchased from Thermo Fisher Scientific (Bedford, MA, USA). Amplex red soluble fluorophore and 4-Methylumbelliferyl phosphate (4-MUP) were purchased from Life Technologies (Carlsbad, CA, USA). Alkaline Phosphatase Streptavidin and the Proton Biotin Protein Labeling Kit were purchased from Vector Laboratories Inc. (Newark, CA, USA). Total and Phospho-Akt/mTOR Pathway panels were purchased from MilliporeSigma (Bedford, MA, USA). The Luminex MAGPIX system was purchased from Luminex Corp. (Austin, TX, USA). The SpectraMax M5 microplate reader was purchased from Molecular Devices Corp. (Sunnyvale, CA, USA).
3
Monoclonal Antibody Generation and Purification
4
Purification and Characterization of ASPH Antibodies
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Pharmaceutical grade ethanol was used in the in vivo experiments. The A85G6 and A85E6 monoclonal antibodies to ASPH and Humbug were generated into human recombinant proteins and purified over Protein G columns (Healthcare, Piscataway, NJ, USA) [34 (link)]. All other primary antibodies used for the duplex ELISAs were purchased from Abcam (Cambridge, MA, USA). RPLPO antibody was sourced from the Proteintech Group Inc (Chicago, IL, USA). ELISA MaxiSorp 96-well plates were purchased from Nunc (Rochester, NY, USA). Horseradish peroxidase (HRP)-conjugated secondary antibody, and Amplex Red soluble fluorophore were purchased from Invitrogen (Carlsbad, CA, USA). The SpectraMax M5 microplate reader was purchased from Molecular Devices Corp. (Sunnyvale, CA, USA). BCA reagents were obtained from Pierce Chemical Corp. (Rockford, IL, USA). All other fine chemicals were purchased from CalBiochem (Carlsbad, CA, USA), Pierce (Rockford, IL, USA), or Sigma (St. Louis, MO, USA).
5
Monoclonal Antibody Production and Purification
6
Monoclonal Antibody Generation and Purification
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Pharmaceutical grade ethanol was used in all experiments. The A85G6 and A85E6 monoclonal antibodies to ASPH were generated to human recombinant protein (de la Monte et al., 2009 (link)) and purified over Protein G columns (Healthcare, Piscataway, NJ). Otherwise, antibodies used for duplex ELISAs were purchased from Abcam (Cambridge, MA). RPLPO) antibody was from the Proteintech Group Inc (Chicago, IL). ELISA MaxiSorp 96-well plates were purchased from Nunc (Rochester, NY). Horseradish peroxidase (HRP)-conjugated secondary antibody, Amplex Red soluble fluorophore, and the Akt Pathway Total and Phospho panels were purchased from Invitrogen (Carlsbad, CA). HRP-labeled polymer conjugated secondary antibody was purchased from Dako Corp (Carpinteria, CA). The SpectraMax M5 microplate reader was purchased from Molecular Devices Corp. (Sunnyvale, CA). BCA reagents were from Pierce Chemical Corp. (Rockford, IL). All other fine chemicals, including NNK were purchased from CalBiochem (Carlsbad, CA), Pierce (Rockford, IL), or Sigma (St. Louis, MO).
7
Monoclonal Antibody Production and Purification
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Pharmaceutical grade ethanol was used in the in vivo experiments. The A85G6 and A85E6 monoclonal antibodies to ASPH were generated to human recombinant protein [46 (link)] and purified over Protein G columns (Healthcare, Piscataway, NJ, USA). Otherwise, antibodies used for duplex ELISAs were purchased from Abcam (Cambridge, MA, USA). RPLPO antibody was from the Proteintech Group Inc. (Chicago, IL, USA). ELISA MaxiSorp 96-well plates were purchased from Nunc (Rochester, NY, USA). Horseradish peroxidase (HRP)-conjugated secondary antibody and Amplex Red soluble fluorophore were purchased from Invitrogen (Carlsbad, CA, USA). The SpectraMax M5 microplate reader was purchased from Molecular Devices Corp. (Sunnyvale, CA, USA). BCA reagents were from Pierce Chemical Corp. (Rockford, IL, USA). All other fine chemicals were purchased from CalBiochem (Carlsbad, CA, USA), Pierce (Rockford, IL, USA) or Sigma (St. Louis, MO, USA).
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