Detection of small molecule diagnostic markers with phage-based open-sandwich immuno-PCR

Open sandwich immunoassay (OS-IA) utilizes antigen-dependent stabilization of antibody variable region to quantify various antigens, enabling noncompetitive detection of small molecules with a broad working range. To further improve its detection sensitivity, here we employed phage-based immuno-PCR approach, wherein OS-IA and quantitative PCR methodologies were combined with the use of immobilized VL fusion protein and filamentous phages displaying VH fragment, whose DNA was extracted for PCR amplification. This approach significantly enhanced the assay sensitivity for small molecule antigens osteocalcin (BGP) peptide and 17beta-estradiol.

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