Flow Immunoassay Using Solid-Phase Entrapment

Abstract

A flow injection immunoassay was performed using a column packed with reversed-phase sorbents to effect separation of the immunoreacted species by entrapping free analyte and allowing antibody-conjugated analyte to pass unretained. Fluorescein-labeled analyte was measured in a competitive assay for the anticonvulsant drug phenytoin. The simplicity of the assay was the greatest advantage of the technique, which allowed for measurement of phenytoin in a 2-min assay time. The reliable detection limit for the assay was 5 nmol L^-1 of phenytoin in serum. The columns were regenerated with periodic injections of ethanol solutions to remove the entrapped analyte and prepare the column for subsequent analyses.