In this work, we compare labeling by two negatively charged fluorescent labels, 8‐aminopyrene‐1,3,6‐trisulfonic acid (APTS) and 8‐(2‐hydrazino‐2‐oxoethoxy)pyrene‐1,3,6‐trisulfonic acid (Cascade Blue hydrazide [CBH]). Effectiveness of the labeling chemistries were investigated by… Click to show full abstract
In this work, we compare labeling by two negatively charged fluorescent labels, 8‐aminopyrene‐1,3,6‐trisulfonic acid (APTS) and 8‐(2‐hydrazino‐2‐oxoethoxy)pyrene‐1,3,6‐trisulfonic acid (Cascade Blue hydrazide [CBH]). Effectiveness of the labeling chemistries were investigated by 4‐hydroxybenzaldehyde and maltoheptaose followed by LC/UV‐MS and CE/LIF analysis, respectively. The reaction yield of APTS labeling was determined to be only ∼10%. This is due to reduction of almost 90% of the analyte by sodium cyanoborohydride to alcohol, which cannot be further labeled via reductive amination. However, the CBH labeling provides ∼90% reaction yield based on the LC/UV‐MS measurements. The significantly higher labeling yield was also confirmed by CE/LIF measurements. Finally, the more effective hydrazone formation technique of CBH was characterized and applied for N‐linked glycan analysis by CE/LIF.
               
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