Time-resolved fluoroimmunoassay for enteric adenoviruses using the europium chelator 4,7-bis(chlorosulfophenyl)-1,10-phenanthroline-2,9-dicarboxylic acid

Abstract

A time-resolved fluoroimmunoassay (TR FIA) was developed for the direct detection of adenovirus types 40 (Ad40) and 41 (Ad41) in stool specimens by using a monoclonal antibody (5D8/2C2) which recognizes both Ad40 and Ad41 but does not cross-react with other adenovirus serotypes. In this assay, the detector antibody is biotinylated directly rather than labeled with europium, and the fluorescent signal is generated on a solid phase in the presence of excess europium (Eu3+). The strength of the signal is dependent on the amount of a Eu3+ chelator [4,7-bis(chlorosulfophenyl)-1,10-phenanthroline-2,9-dicarboxylic acid (BCPDA)]-streptavidin complex bound by the biotinylated detector antibody (5D8/2C2). In a pilot study with 41 specimens, this TR FIA demonstrated a maximum sensitivity and specificity of 88% compared with SmaI restriction analysis of adenovirus isolates from the same specimens. TR FIA using the europium chelator BCPDA represents a feasible approach for the direct identification of specific adenovirus serotypes in stool specimens.