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Chicken Infectious Anaemia Virus | 255

Figure 9.4 Maps of two sets of chimeras with substitutions of VP1 coding regions. The ORF map at the top of the figure shows the three
CIAV ORFs, and lowercase single-letter amino acid codes indicate the amino acid differences found in CIA-1 compared with Cux-1(C).
Positions of the restriction sites in relation to the amino acid differences are indicated above the VP1 box. Renshaw et al., Journal of Virology,
Vol. 70, No. 12 Dec. 1996, pp. 8872–8878. Copyright @ 1996, with permission of the American Society for Microbiology.

Figure 9.5 Percentages of infected cells at time points following transfection with Cux-1(C), CIA-1, and the chimeras depicted in Fig.
6
9.4. Cells (2 × 10 ) were transfected with 500 ng of each ligation product and 500 ng of pRC/RSV-lacZ. Cells were examined at 24 hours
post transfection for β-galactosidase production to verify the relative consistency of the transfections (data not shown). The presence
of VP3 was verified initially at 48 hours post transfection and at various time points thereafter. Non-infect. Clone = defective CIA-1 clone.
IFA+ = immunofluorescence assay positive. Renshaw et al., Journal of Virology, Vol. 70, No. 12 Dec. 1996, pp. 8872–8878. Copyright @ 1996,
with permission of the American Society for Microbiology

and secondary reagents is essential, because false positive stain- al., 2002; van Santen et al., 2004b). Early detection of virus
ing has been described using PBS as the primary reagent and replication can also be achieved by using qRT-PCR assays
FITC- or HRPO-labelled goat, rabbit or horse anti-chicken (Markowski-Grimsrud et al., 2002). A detailed description for
IgG as the secondary reagent (Lucio et al., 1991). Currently, virus isolation, identification, and propagation is provided by
real-time quantitative (q)PCR assays are preferred to dem- Smyth and Schat (2016).
onstrate an increase of virus in supernatant fluids and several In addition to the susceptible cell lines, mononuclear spleen,
qPCR protocols have been described (Markowski-Grimsrud et thymus, and bone marrow cells can be infected with at least