FIG. 5.

DNA laddering and NP and aNP patterns in chicken eggs. Seven-day-old chicken embryos were infected in the allantoic cavity with WT and delNS1 viruses at an MOI of 20 to 100 PFU per egg. (A) At 48 hpi, samples of allantoic fluid were taken from four eggs separately, and DNA was isolated by SDS-high-salt extraction of 150 μl of allantoic fluid from each egg and analyzed by electrophoresis on a 1.6% agarose gel. Lane M, 0.4 μg of MassRuler DNA ladder mix (MBI Fermentas). (B) Samples of allantoic fluid were taken from five eggs separately and NP and aNP patterns were analyzed by PAGE-WB. (C) Allantoic fluid samples from two separate eggs were sedimented through a 40% glycerol cushion at 100,000 × g for 2.5 h. Equivalent amounts of virus pellet (VP) and supernatant (S) were analyzed by PAGE-WB. NP and aNP were identified by ECL with an anti-NP mouse antibody (clone A3) and anti-mouse HRP conjugate.