Fluorescent Nanoparticle-Based Indirect Immunofluorescence Microscopy for Detection ofMycobacterium tuberculosis

Abstract

A method of fluorescent nanoparticle-based indirect immunofluorescence microscopy (FNP-IIFM) was developed for the rapid detection ofMycobacterium tuberculosis. An anti-Mycobacterium tuberculosisantibody was used as primary antibody to recognizeMycobacterium tuberculosis, and then an antibody binding protein (Protein A) labeled with Tris(2,2-bipyridyl)dichlororuthenium(II) hexahydrate (RuBpy)-doped silica nanoparticles was used to generate fluorescent signal for microscopic examination. Prior to the detection, Protein A was immobilized on RuBpy-doped silica nanoparticles with a coverage of∼5.1×102molecules/nanoparticle. With this method,Mycobacterium tuberculosisin bacterial mixture as well as in spiked sputum was detected. The use of the fluorescent nanoparticles reveals amplified signal intensity and higher photostability than the direct use of conventional fluorescent dye as label. Our preliminary studies have demonstrated the potential application of the FNP-IIFM method for rapid detection ofMycobacterium tuberculosisin clinical samples.