Cy5 hydrazide is a reactive dye which reacts smoothly and nearly quantitatively with various carbonyl groups encountered in biomolecules. The labeling regent is used for the labeling of aldehydes and ketones. The dye examples are proteins subjected to oxidative stress, glycoproteins pre-activated by periodate oxidation, and oligonucleotides with aldehyde moieties. Cy5 hydrazide could replace carbonyl-reactive Alexa Fluor 647 and DyLight 649 dyes. For biomolecule labeling, the labeling regent has low aqueous solubility, using of organic co-solvent to dissolve this molecular is necessary for efficient reaction. First, Cyanine dye should be dissolved in organic solvent and then added to a solution of biomolecule in appropriate aqueous buffer.
In HL-60 cells, Protein carbonylation induced by Hydrogen peroxide (H2O2) was quantitatively analyzed by using fluorescent dyes of Cy5-hydrazide dye, followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and fluorescence determination [1].
Reference:
[1] Yoshimura, T. ; Harashima, M.; Kurogi, K.; Suiko, M.; Liu, M.-C.; Sakakibara, Y. A novel procedure for the assessment of the antioxidant capacity of food components. Analytical Biochemistry, 2016, 507, 7–12.