ABSTRACT
Frontal affi nity chromatography (FAC) is a quantitative technique of affi nity chromatography originally developed by Kasai and Ishii [1] (for a detailed review, see Ref. [2]). Although it was fi rst investigated as a tool for biomolecular interactions between enzymes and their inhibitors, its high potential and versatility have been demonstrated for many other biomolecules, for example, for quantitative specifi city analysis of a representative plant lectin, concanavalin A, using a series of radiolabeled asparagine-linked oligosaccharides [3]. Originally, FAC was operated manually in a cold room using an open column and a fraction collector. Such procedures are laborious, time-consuming, and not suffi ciently reproducible. In 2000, the system was greatly improved by the incorporation of a high-performance liquid chromatography (HPLC) system to assure high reproducibility and by the introduction of fl uorescently labeled oligosaccharides to achieve high sensitivity [4]. The fi rst such system introduced was quite simple, comprising a single isocratic pump, a manual injector, a fl uorescence detector, and a personal computer. A specialized data analysis program was also developed [5]. With this specialized system, several dozens of interaction
analyses per day were performed for the determination of dissociation and association constants (Kd and Ka, respectively) between galectins and pyridylaminated (PA) oligosaccharides [6,7].
