Azo biotin azide

For the acid-cleavable reagent (DADPS Biotin–Azide, CLICK CHEMISTRY TOOLS)-ligated sample, streptavidin-bound peptides were eluted using 10% of formic acid (FA) three times at room temperature. For the reduction-cleavable reagent (Azo Biotin–Azide, Sigma-Aldrich)-ligated sample, streptavidin-bound peptides were eluted using 300 mM Na₂S₂O₄ in 20 mM HEPES, 6 M urea, and 2 M thiourea buffer, pH 7.6, at room temperature. For the photo-cleavable reagent (UV Cleavable Biotin–Azide, Kerafast)-ligated sample, streptavidin-bound peptides were eluted by exposing under 365 nm UV light for 1 h at room temperature. The peptides were collected and desalted with home-made C18 Tips.
Before peptide release, the photo-cleavable reagent labeled sample should be performed with light protection, and the acid-cleavable reagent labeled sample should be performed avoiding acid and not doing the SCX procedure.