Fig. 4.

Virus susceptibility of the continuous porcine myelomonocytic cell lines 3D4/2, 3D4/21 and 3D4/31 in 7.5% FBS/RPMI, compare to the susceptibility of standard assay cells: 3D4/2 (dotted column), 3D4/21 (white), 3D4/31 (dashed), standard assay cells (black column). Panel A illustrates results when cells were seeded in 96-well plates and incubated overnight in RPMI containing 15% FBS. The maintenance medium was then replaced with 10-fold dilutions of various virus stocks made in RPMI and incubated with the cells for 3 h at 37 °C, 5% CO₂. After the adsorption period, the virus inoculum was removed and cells were washed once with RPMI and overlayed with their respective maintenance medium containing 7.5% of FBS. Cells were incubated for 5 days at 37 °C, 5% CO₂ and then either fixed for immunostaining (ASFV, CSFV) or evaluated for the degree of cytopathic effect (CPE) (SVDV, VSV, PRV). In the case of cells infected with SwPV, the infected cell supernatant was titrated using a susceptible cell line (ST). Panel B illustrates results when viruses were successively passed twice in each of the three cell lines and the supernatants from the second passage were then titrated (10-fold dilutions) on previously determined, susceptible cell lines along with the original virus inoculum. Infection of the IPAM cell clones was carried out by incubating a 0.5 ml cell suspension (2×10⁵ cells/ml) made in RPMI containing 7.5% FBS with 0.5 ml of virus inoculum in a 12-well plate for 5 days at 37 °C, 5% CO₂. Prior to preparing the cell suspension, the IPAM cell clones were maintained in the above medium containing 15% FBS.