Cysteine supplementation further increases FL-gB expression in infected High Five.

<p>(<b>A</b>) 300x concentrated cysteine supplement was prepared in culture medium to obtain the indicated final Cys concentrations and added alone to High Five cell cultures 24 h.p.i. with bBst2x at CCI 2.5 with m.o.i. 5. Cys was added alone (left panel) or in combination with 50 nM rapamycin treatment (right panel). Cells were harvested at 72 h.p.i. and detergent-soluble FL-gB relative quantification performed as already described (controls were made by adding fresh medium to infected cell cultures previously supplemented or not with 0.1% DMSO at the time of infection). (<b>B</b>) FL-gB expression was compared in detergent-soluble (Sol) or -insoluble (Ins) protein extracts obtained from cells treated as in (A right panel). Equal loading was by resuspending the insoluble pellets in the same volume used to prepare the soluble fractions. Immunoblot (left panel) and densitometric analysis (right panel) are shown. (<b>C</b>) Effect on the dissolved oxygen (DO) control and cell viability in bioreactors of bBst2x-infected High Five - CCI 2.5, m.o.i. 5, rapamycin 50 nM - by either cysteine shot addition (Shot, dashed lines and triangles) or continuous feeding (Cont, solid lines and circles). Bioreactors were sampled for cell viability, while DO was recorded in real-time and expressed as the percentage of air saturation.