Optimization of a FACS based-immunoassay using E. coli autodisplaying Z-domains

Abstract

We developed a fluorescence-activated cell sorter (FACS)-based immunoassay using Escherichia coli with autodisplayed Z-domains as a solid support for detection antibodies as well as a method to optimize the signal-to-noise ratio of fluorescence signals from E. coli. To improve the sensitivity of FACS measurements by eliminating artifactual signals from unrelated small particles in the sample mixture, E. coli with autodisplayed Z-domains was modified to express enhanced green fluorescence protein (eGFP) in the cytosol. By selectively gating the measurement channel for eGFP in the cytosol, we were able to discriminate between green fluorescent E. coli and artifact particles. We confirmed the efficacy of this optimization method for FACS-based immunoassay using several model reactions. Finally, as proof of concept, we used the FACS-based immunoassay and our optimization method for the medical diagnosis of human hepatitis B virus surface antigen.