Amplified fragments of HMPV DNA derived from the Veto cell
culture supernatant were subjected to direct sequencing.
Comparison of Veto cell plaque assay, TaqMan reverse transcriptase polymerase chain reaction RNA assay, and VecTest antigen assay for detection of West Nile virus in field-collected mosquitoes.
To assess virus viability in grinding solution, 50 [micro]L of each tested grinding solution and virus dilution was placed into 450 [micro]L of BA-1 diluent, and aliquots of 200 [micro]L of this mixture were added to Veto cells reared on standard nutrient medium (i.e., Dulbeeco's modified Eagle medium and 10% fetal bovine serum [FBS]) at 37[degrees]C and 5% [CO.sub.2],.
Negative antigen (uninfected Veto cell lysate produced as described above) was placed in every third column of the plate (i.e., columns 1, 4, 7, 10), and positive antigen was placed in the remaining columns.
In the morning, 100 [micro]L of each serum-virus mixture was added onto confluent monolayers of Veto cells and allowed to adsorb at 37[degrees]C for 1 h.
conorii Malish #7 grown in Veto cells
) and negative controls that were extracted and PCR amplified in parallel with the specimens.
A total of 7 urine samples (all from the third visit) produced a characteristic CPE on Veto cells
after 5 to 7 days of culture.
Virus stocks were prepared in Veto cells
, and each animal was inoculated with approximately 1,000 PFU.
The SARSCoV (strain HKU-39849) used in both VNT and IFA was plaque purified three times in Veto cells
, and stock virus (titer 5 x [10.sup.7] 50% tissue culture infective dose [TCI[D.sub.50]]) prepared by two low-multiplicity passes in Veto cells
The antigen used was a precipitate (polyethylene glycol 6000) of the supernatant of Veto cells
infected with the RVFV clone 13.
Isolation of SARS-CoV was performed with Veto cells
maintained in Dulbecco's modified Eagle medium (D-MEM, BioWhittaker, Verviers, Belgium) and supplemented with 10% fetal calf serum (FCS, HyClone, Perbio Science Erembodegem-Aalst, Belgium), penicillin/streptomycin (BioWhittaker), and 2.5 [micro]g/mL Fungizone (Invitrogen Ltd, Life Technologies, Paisley, UK) (complete medium).
In 2001, fathead minnow (FHM), white sturgeon skin (WSS), epithelioma papillosum caprini (EPC), and channel catfish ovary (CCO) cells were used instead of the Veto cells
. Inoculated cells were incubated in a 5% C[O.sub.2] atmosphere at 37[degrees]C.