Cy5 alkyne, with low aqueous solubility is a dye ready for the use in Click Chemistry reaction. Various molecules via Click Chemistry reaction can be attached by this deeply colored, and photostable Cy5 fluorophore. Most derivatives of non-sulfonated cyanines have low aqueous solubility except for hydrochlorides of hydrazides and amines. For biomolecule labeling, 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. Various substrates such as biomolecules, polymers, and solid surfaces which bearing azides can be used for the labeling.
In the azido-GalNAc samples, alkyne-Cy3 and alkyne-Cy5 specifically label azido-proteins. Furthermore, researches also indicate that the Cy5 and Cy3 probe-protein conjugates have non-overlapping fluorescence profiles and can be differentially detected within a single gel [1].
Reference:
[1] Burnham-Marusich, A. R.; Plechaty, A.M.; Berninsone, P.M. Size-matched alkyne-conjugated cyanine fluorophores to identify differences in protein glycosylation. Electrophoresis, 2014, 35(18), 2621-2625.