ALTEX :: Alternatives to Animal Experiments

1993, Volume 1

Quantification of Lymphocytic ChorioMeningitis Virus with an Immunological Focus Assay in 24 Well Plates

Manuel Battegay

Institut fuer Experimentelle Immunologie, Departement Pathologie der Universitaet CH-Zurich

SUMMARY

Titres of lymphocytic choriomeningitis virus (LCMV) were determined on adherent fibroblast cell lines in 24, respectively 96 well plates. After absorption of virus by cells and 48 h incubation under a methylcellulose overlay, cell monolayers were fixed with 4% formaldehyde in phosphate buffered saline, permeabilized by incubating in 0.5% Triton X-100 in balanced salt solution and then stained with a monoclonal rat anti-LCMV and a peroxidase labeled second stage antibody. The sensitivity of the assay is within a factor of 2-4 of conventional plaquing methods. The method is quicker (2-3 days), as compared to conventional methods (4-6 days) and less expensive with respect to both workhours and materials involved. The method also detects poorly- or non-plaquin LCMV isolates, and therefore drastically reduces the needs for titration of LCMV in mice.

Keywords: lymphocytic choriomeningitis virus, quantification in vitro, plaquing method