Labeling a protein with fluorophores using NHS ester derivitization
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Updated: May 4, 2026

Proteome-wide Quantification of Labeling Homogeneity at the Single Molecule Level
Published on: April 19, 2019
Jagpreet S Nanda1, Jon R Lorsch1
1Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
Abstract:
N-hydroxysuccinimde (NHS) ester-mediated derivitization involves the reaction of this amine-reactive group with the primary amines of a protein or a biomolecule. Using NHS chemistry allows one to conjugate various fluorescent probes, biotin, and cross-linkers to primary amines. For example, we use NHS ester chemistry to fluorescently label the amino terminus of a protein with the dye, 5-(and-6)-carboxyfluorescein, succinimidyl ester (5(6)-FAM, SE).
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