Propagation of hepatitis A virus in a renal cell line JTC-12.P3 of cynomolgus monkey origin.
Human hepatitis A virus (HAV) derived from 10% HAV infected marmoset liver homogenate and faeces from acute hepatitis A was successfully propagated in vitro in a new cell line, JTC-12.P3. The cell line originated from the renal cortex of cynomolgus monkey which was adapted to growth in a serum free, protein free, chemically defined synthetic medium. Replication of the virus was followed by solid phase RIA, immunofluorescent staining, and immunoelectron microscopy. The propagation of HAV occurred over several passages, with the 1st and 2nd passages requiring at least 8 weeks each. However, with the increasing serial passage of virus, the period needed to detect it was shortened, suggesting the adaptation of HAV to the cells. The identity of the newly synthetized virus particles with HAV was established by immunoelectron microscopy and immunofluorescent blocking effect with human convalescent serum. The HAV propagated in JTC-12.P3 cells banded predominantly at a density of 1.32 g/cm3 in CsC1 gradient. The infected cells showed no specific signs of CPE. Ultrastructurally, clusters of virus particles 27 nm in diameter were observed mainly in the lysosomal vesicles and freely in crystalline array in the cytoplasm, too. Addition of 0.1% of various anti-HAV negative sera or of prostaglandin E1 to the culture medium caused accelerated propagation of HAV.
Baba
M
M
Third Department of Internal Medicine, Mie University School of Medicine, Japan.
Takegawa
M
M
Kaito
M
M
Miyamoto
K
K
Suzuki
S
S
D000818:Animals; D000956:Antigens, Viral / Q000032:analysis; D002460:Cell Line / Q000382:microbiology*; D002500:Centrifugation, Isopycnic; D006507:Hepatovirus / Q000254:growth & development*; D007672:Kidney Cortex / Q000166:cytology; D008252:Macaca fascicularis; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D012680:Sensitivity and Specificity; D014776:Virus Cultivation / Q000379:methods*
D000956:Antigens, Viral