Investigating noncovalent squarylium dye–protein interactions by capillary electrophoresis–frontal analysis

Noncovalent interactions between fluorescent probe molecules and protein analyte molecules, which typically occur with great speed and minimal sample handling, form the basis of many high sensitivity analytical techniques. Understanding the nature of these interactions and the composition of the resulting complexes represents an important area of study that can be facilitated by capillary electrophoresis (CE). Specifically, we will present how frontal analysis (FA) and Hummel-Dreyer (HD) methods can be implemented with CE to determine association constants and stoichiometries of noncovalent complexes of the red luminescent squarylium dye Red-1c with bovine serum albumin (BSA) and β-lactoglobulin A. By adjusting solution conditions, such as pH or ionic strength, it is possible to selectively modify the binding process. As such, conditions for optimal selectivity for labeling reactions can be established by capillary electrophoresis–frontal analysis (CE–FA) investigations.