THEORETICAL CONCEPTS OF ONLINE LIQUID-CHROMATOGRAPHIC BIOCHEMICAL DETECTION SYSTEMS .2. DETECTION SYSTEMS BASED ON LABELED AFFINITY PROTEINS

A theoretical concept for on-line liquid chromatography-biochemical de tection (LC-BCD) using labelled affinity proteins as reporter molecule s is presented. The BCD system is based on the post-column addition of labelled affinity proteins such as fluorescein-labelled streptavidin to the LC effluent. After a short reaction time, free and analyte-boun d label are separated during passage through a column packed with an i mmobilised-ligand support. The bound fraction passes the column unreta ined and is measured downstream by means of a conventional HPLC detect or. The theoretical model presented here relates the detector response to the most important instrumental and biochemical parameters such as dispersion, reaction time, concentration and affinity of the affinity protein and the number of binding sites. The theoretical concept is v alidated using fluorescein-labelled streptavidin and biotin as model s ystem. (C) 1997 Elsevier Science B.V.