Indocyanine green as a noncovalent, pseudofluorogenic label for protein determination by capillary electrophoresis

Indocyanine green (ICG)--a negatively charged, polymethine dye--can interact noncovalently with proteins to form fluorescent complexes, with excitation and emission maxima near 780 and 820 nm, respectively. This behavior was realized utilizing either a 100 mM phosphate buffer or a 25 mM citric acid buffer, both at pH 3.1. The behavior of ICG under these conditions, termed pseudofluorogenic, rendered the dye suitable for use as a label for protein determination in capillary electrophoresis with diode laser-induced fluorescence detection (CE-LIF). To this end, pseudofluorogenic ICG was used both as an on-column label for human serum albumin (HSA) and as a precolumn label for a model mixture of proteins, including ribonuclease A, transferrin, and cytochrome c. These ICG-labeled proteins were successfully resolved in less than 11 min, with no interference from excess, unbound dye.