Figure 6

Density analysis of VLPs released from transfected cells expressing NiV proteins. (A) Clarified culture supernatants from cells expressing different combinations of NiV proteins were layered onto 5–45% sucrose gradients and centrifuged. Fractions were immunoprecipitated with MAbs F45G6 (ant-N), F45G5 (anti-M) or rabbit polyclonal anti-serum against NiV F or HeV G. A portion of each fraction was used to determine density. Densitometry analysis of the gels shown in (A) was used to determine the quantitative distribution of proteins (B). Where more than one protein was expressed, the protein analyzed by densitometry is indicated by parentheses. (C) Supernatant from cells infected with NiV was layered onto a 5–45% sucrose gradient and centrifuged. RNA was extracted from fractions and used for real-time PCR. The threshold cycle (Ct) value was used as a proxy for virus concentration.