Evaluation of a Japanese quail fibrosarcoma cell line (QT-35) for use in the propagation and detection of metapneumovirus.
A Japanese quail fibrosarcoma cell line (QT-35) was evaluated and compared to Vero cells for its utility in metapneumovirus propagation, titration and serological detection by indirect immunofluorescence staining. Cell characteristics such as growth kinetics at different passage levels and seeding density in 96-well plates using various media formulations were studied in order to determine suitable assay parameters. Specifically, QT-35 cells supported the replication of a subgroup A metapneumovirus, strain 14/1, when maintained in DMEM containing a high level of glucose (4500 mg/l) and 2% gamma-irradiated fetal bovine serum (gamma-FBS). There appeared to be a decreased ability of metapneumovirus produced in chicken embryo fibroblast (CEF) cells to replicate to high titers in QT-35 cells, however, this apparent restriction was overcome after the second passage resulting in high titered stock. Metapneumovirus produced in Vero cells and propagated in QT-35 cells produced high titered stock after the first passage. Viral titers determined in Vero and QT-35 cells were comparable, when the latter cell line was used at passage levels < or = 20 and seeded between 5.0 x 10(4) and 1.0 x 10(5) cells/0.33 cm(2) in hgDMEM containing 10% gamma-FBS, with a reduction to 2% gamma-FBS when the virus was applied to the cell monolayers 24 h post-seeding. After infection with metapneumovirus, QT-35 cells exhibited syncytia, similar to those in metapneumovirus-infected Vero cells, which were readily detected by indirect immunofluorescent (IF) staining.
Sabara
Marta I
MI
National Centre for Foreign Animal Diseases, The Canadian Science Centre for Human and Animal Health, 1015 Arlington Street, MB, R3E 3M4, Winnipeg, Canada. msabara@inspection.gc.ca
Larence
June E
JE
D000818:Animals; D000914:Antibodies, Viral / Q000032:analysis; D002455:Cell Division; D002522:Chlorocebus aethiops; D003370:Coturnix; D005354:Fibrosarcoma; D019084:Fluorescent Antibody Technique, Indirect; D029121:Metapneumovirus / Q000276:immunology / Q000302:isolation & purification / Q000502:physiology*; D013194:Staining and Labeling / Q000379:methods; D017186:Titrimetry; D014407:Tumor Cells, Cultured; D014709:Vero Cells; D014779:Virus Replication
D000914:Antibodies, Viral