Cell culture
Chicken macrophage-like cell line HD-11 (Beug et al., 1979) was
cultivated at 37 C, 5% CO2 in complete RPMI-1640 (RPMI-1640
supplemented with 10% heat-inactivated foetal calf serum (HI-FCS),
1 GlutaMAX, 50 U/ml penicillin, 50 mg/ml streptomycin (all reagents
from Life Technologies)). For maintenance and long incubation
during knock-down experiments, heat inactivated chicken
serum (Life Technologies) was added to 1% final concentration.
Bone marrow was obtained from the femurs of new-born male
chicks, under approval from the University of Queensland animal
ethics committee. Chicken bone marrow-derived macrophages
(BMMs) were obtained by cultivation of marrow for one week in
complete RPMI-1640 supplemented with 15% conditioned medium
from CHO cells transiently expressing chicken CSF1 (Garceau et al.,
2010), generated as follows. CHO suspension cells were grown in
CD CHO medium (Gibco) supplemented with 8 mM glutamine
(Gibco).