Concentration of hydroxyl radicals during electrical discharge for radical probe protein footprinting mass spectrometry

Hydroxyl Radical Protein Footprinting: A Mass Spectrometry-Based Structural Method for Studying the Higher Order Structure of Proteins

“…Noteworthy, Downard also demonstrated the functionality of optical dosimeters in determining the concentration of hydroxyl radicals in the context of electrospray discharge experiments . Indigo carmine typically has a maximum absorption at 610 nm; however, reaction with hydroxyl radicals causes a loss of conjugation, resulting in a loss of the absorbance band.…”

“…Indigo carmine typically has a maximum absorption at 610 nm; however, reaction with hydroxyl radicals causes a loss of conjugation, resulting in a loss of the absorbance band. To evaluate the efficacy of indigo carmine to act as a dosimeter, sample solutions containing hen egg white lysozyme and 1 mM indigo carmine were subjected to oxidation via RP-MS; 20 μL of the resulting condensate was collected and diluted to 100 μL with water and transferred to a 96-well plate which was analyzed using a multiplate reader . By decreasing the flow rate of the ESI syringe, it was possible to increase the exposure time to the hydroxyl radicals.…”

“…Noteworthy, Downard also demonstrated the functionality of optical dosimeters in determining the concentration of hydroxyl radicals in the context of electrospray discharge experiments. 84 Indigo carmine typically has a maximum absorption at 610 nm; however, reaction with hydroxyl radicals causes a loss of conjugation, resulting in a loss of the absorbance band. To evaluate the efficacy of indigo carmine to act as a dosimeter, sample solutions containing hen egg white lysozyme and 1 mM indigo carmine were subjected to oxidation via RP-MS; 20 μL of the resulting condensate was collected and diluted to 100 μL with water and transferred to a 96-well plate which was analyzed using a multiplate reader.…”

“…To evaluate the efficacy of indigo carmine to act as a dosimeter, sample solutions containing hen egg white lysozyme and 1 mM indigo carmine were subjected to oxidation via RP-MS; 20 μL of the resulting condensate was collected and diluted to 100 μL with water and transferred to a 96-well plate which was analyzed using a multiplate reader. 84 By decreasing the flow rate of the ESI syringe, it was possible to increase the exposure time to the hydroxyl radicals. Plotting indigo carmine absorbance as a function of increasing exposure time resulted in a linear decrease in signal corresponding to degradation of the optical dye.…”