title: Accelerated and improved quantification of lymphocytic choriomeningitis virus (LCMV) titers by flow cytometry.
abstract: Lymphocytic choriomeningitis virus (LCMV), a natural murine pathogen, is a member of the Arenavirus family, may cause atypical meningitis in humans, and has been utilized extensively as a model pathogen for the study of virus-induced disease and immune responses. Historically, viral titers have been quantified by a standard plaque assay, but for non-cytopathic viruses including LCMV this requires lengthy incubation, so results cannot be obtained rapidly. Additionally, due to specific technical constraints of the plaque assay including the visual detection format, it has an element of subjectivity along with limited sensitivity. In this study, we describe the development of a FACS-based assay that utilizes detection of LCMV nucleoprotein (NP) expression in infected cells to determine viral titers, and that exhibits several advantages over the standard plaque assay. We show that the LCMV-NP FACS assay is an objective and reproducible detection method that requires smaller sample volumes, exhibits a ∼20-fold increase in sensitivity to and produces results three times faster than the plaque assay. Importantly, when applied to models of acute and chronic LCMV infection, the LCMV-NP FACS assay revealed the presence of infectious virus in samples that were determined to be negative by plaque assay. Therefore, this technique represents an accelerated, enhanced and objective alternative method for detection of infectious LCMV that is amenable to adaptation for other viral infections as well as high throughput diagnostic platforms.
authors.affiliation: Integrated Department of Immunology, University of Colorado Denver and National Jewish Health, Denver, Colorado, United States of America. darlynn.kornsjohnson@ucdenver.edu
authors.forename: Darlynn
authors.initials: D
authors.lastname: Korns Johnson
authors.forename: Dirk
authors.initials: D
authors.lastname: Homann
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000096:biosynthesis; D005434:Flow Cytometry / Q000379:methods*; D008216:Lymphocytic Choriomeningitis / Q000097:blood / Q000276:immunology / Q000821:virology*; D008217:Lymphocytic choriomeningitis virus / Q000254:growth & development* / Q000378:metabolism; D051379:Mice; D009698:Nucleoproteins / Q000096:biosynthesis* / Q000276:immunology; D012680:Sensitivity and Specificity; D019562:Viral Load / Q000379:methods*; D010948:Viral Plaque Assay; D014766:Viremia / Q000175:diagnosis
chemical_list: D000914:Antibodies, Viral; D009698:Nucleoproteins