title: Biochemical detection for direct bead surface analysis.
abstract: A continuous-flow biochemical detection system is presented which recognizes biologically active compounds immobilized to solid phases. This approach can be used to screen, for example, solid-phase combinatorial libraries for lead compounds. Biochemical detection is performed by mixing a plug of a solid-phase suspension with labeled affinity protein. During a short reaction time, the labeled affinity protein will only bind to ligands, i.e., compounds with biological activity. Hereafter, the free and bound labels are separated by means of a hollow fiber module. Quantitation of the free label is performed with a conventional flow-through fluorescence detector. Total assay time amounts to less than 3 min. Biochemical detection for direct bead surface analysis was developed for two model systems. The first model system used fluorescence-labeled avidin as affinity protein and its ligands biotin and iminobiotin immobilized to agarose as analytes. The second model system used fluorescence-labeled antisheep (Fab)(2) fragments as affinity protein and different IgGs immobilized to agarose as analytes. The feasibility of this approach for recognition of solid-phase immobilized ligands was documented by screening 50 samples with a 100% hit rate.
authors.affiliation: Division of Analytical Chemistry, Leiden/Amsterdam Center for Drug Research, Leiden University, P.O. Box 9502, 2300 RA Leiden, The Netherlands.
authors.forename: E S
authors.initials: ES
authors.lastname: Lutz
authors.forename: H
authors.initials: H
authors.lastname: Irth
authors.forename: U R
authors.initials: UR
authors.lastname: Tjaden
authors.forename: J
authors.initials: J
authors.lastname: van der Greef