Multiplexed Flow Cytometric Immunoassay for Influenza Virus Detection and Differentiation

Abstract

Microsphere-based immunoassay by flow cytometry has gained popularity lately in protein detection and infectious disease diagnosis due to its capacity for multiplexed analysis and simple assay format. Here, we demonstrated the power of microsphere-based immunoassay for high-sensitivity detection and accurate differentiation of influenza viruses. The effects of sample volume and bead number on the assay sensitivity of viral antigen detection were studied. Compared to enzyme-linked immunosorbent assays, flow-based bead assays provided approximately 10-fold lower detection limit for viral particle detection and performed similarly for recombinant viral hemagglutinin protein detection. A four-plexed assay for influenza virus typing and influenza B virus sublineage characterization was developed to demonstrate the potential for multiplexed viral antigen detection and differentiation.

Nanoparticle-based flow virometry for the analysis of individual virions

“…Up to now, the presence of these proteins has been studied in viral preparations either with ELISA or with flow cytometry of multiple viruses bound to microbeads (9,12,(22)(23)(24)(25)(26)(27). These studies analyzed viral particles in bulk.…”

Approaching clinical proteomics: current state and future fields of application in fluid proteomics

“…Thus, up to 100 different sets with each matched to a different probe molecule can be used (97,98). Such systems have been used to determine the concentration of cytokines (97,99), screening for cystic fibrosis (97, 100), hepatitis B sero conversion and human immunodeficiency virus (97,101,102), thyroid hormones (97,103), kinase testing (97,104), allergy testing (97,105), single nucleotide polymorphisms (97,106), infectious disease diagnosis (97,107), and detection of biological warfare agents (97,108) or antibodies in serum or cell culture supernatant (109,110).…”

Multiplexed Immunoassays

“…The features of these xMAP systems are listed in Table 3. The xMAP technology has been demonstrated as a powerful multiplexing method and has found applications in detection of human cytokines 40 , single nucleotide polymorphisms (SNPs) 41 , allergy testing 42 , infectious disease diagnosis 43 , and biological warfare agent screening 44 . Another similar optically encoded microsphere based multiplexing platform was developed by Becton Dickinson BioSciences, named Cytometric Bead Array system (BD TM CBA).…”