A Practical Cytopathic Effect/Dye-Uptake Interferon Assay for Routine Use in the Clinical Laboratory

Disturbance of Tumor Necrosis Factor Alpha-Mediated Beta Interferon Signaling in Cervical Carcinoma Cells

“…These data illustrate that the antiviral activity of TNF-␣ was reconstituted in nonmalignant hybrids even in the presence of the viral oncoproteins but was lost in tumorigenic segregants or parental HeLa cells. To estimate how much IFN-␤ was definitively synthesized, the supernatants of nonmalignant hybrids were removed and tested for IFN-␤ activity against EMCV on A549 indicator cells, where TNF-␣ itself had no antiviral activity (66). Concordant with the RT-PCR data (Fig.…”

“…To verify that the IFN-␤-specific mRNA was translated into functional protein, a biological assay based on the susceptibility of nonresponsive cells to lysis after infection with EMCV was used (37,66) (Fig. 2).…”

“…The cervical carcinoma cell lines HeLa, SW756, CaSki, and SiHa; the nontumorigenic somatic cell hybrids made between HeLa cells and normal human fibroblasts (444 cells) and their tumorigenic segregants (CGL3 cells) (61); HeLa ϫ CaSki and HeLa ϫ SW756 hybrids (15,59); SiHa ϫ HeLa hybrids (see below); malignant HPK Ia cells (10); and the human lung carcinoma cell line A549 (66) were maintained in Dulbecco's modified Eagle's medium supplemented with 10% fetal calf serum and 1% penicillin-streptomycin. Modified SiHa cells (so-called universal fuser) (50) harboring dominant (G418 resistance) and recessive (hypoxanthine-phosphoribosyltransferase-negative) phenotypes were grown in the same medium containing 1 mg of G418 (Gibco BRL) per ml and 10 Ϫ4 M 6-thioguanine (Sigma).…”

“…Antiviral titration assays. Cells were infected with either EMCV or VSV using multiplicities of infections (MOIs) of between 0.1 and 0.2 (37,66). The cells, seeded in 96-well plates (10 4 cells/well) were pretreated with cytokines as described in the figure legends.…”

Differential Antiviral Response of Endothelial Cells after Infection with Pathogenic and Nonpathogenic Hantaviruses

“…HUVECs were maintained in MCDB131 (Gibco BRL, Karlsruhe, Germany) supplemented with 10% fetal calf serum, 100 IU of penicillin/ml, 100 g of streptomycin/ml, 1 g of amphotericin B/ml, 4.5 mM L-glutamine, and 20 g of endothelial-cell growth factor/ml. Vero E6 cells and A549 cells (48) were maintained in Dulbecco's modified Eagle's medium supplemented with 10% fetal calf serum, 100 IU of penicillin, 100 g of streptomycin/ml, and 4.5 mM L-glutamine. The medium and fetal calf serum were certified endotoxin free by the manufacturers.…”