Cells were washed in PBS and incubated with 5 mM CFSE or 5 µg/ml CMPTX (Molecular Probes) in OPTIMEM medium (Life Technologies), at 37°C, 5% CO2, for 20 min. Then cells were thoroughly washed with PBS and cultured for a supplementary hour at 37°C before mixing with PBMC, or overnight for mixing between CFSE- and CMPTX-stained cells.
She is on supplementary oxygen 24 hours a day and spends most of her time in hospital having strong antibiotics delivered into her bloodstream, to help keep her alive while she waits for her transplant.
TUNEL-positive cells were noted to extend to the midbrain of embryos treated with nestin MO for 49 hours (Supplementary Fig. S2).
To determine the time dependence of the drug effect of mitomycin C, cells were treated with desferrioxamine, which caused a time-dependent increase in HIF-1α protein expression with a maximal effect at approximately 10 hours (Supplementary Fig. S2c).
Instead, GFP-expressing Photorhabdus appeared to be cleared from the gut within 24 hours (supplementary material Fig. S7).
Sma expression increased during development, with a peak level at about 12 hours (supplementary material Fig. S7).
TiPM were stimulated with muramyl dipeptide (MDP), Pam3 and HkEc for 24 hours (supplementary material Fig. S5E).
IL1β was induced by short-term fasting but returned to normal at 48 hours (Supplementary Figure 1D), when hepcidin mRNA expression was still increased markedly.
A similar effect of CXCR4 inhibition on EGFR expression was observed in GDF15 but not in FGF2-treated cultures after 24 hours (supplementary material Fig. S3D).
However, these worms were not irreversibly damaged because they recovered their ATP production when transferred to in vitro culture with the colonic LS174T cell line for 24 hours (Supplementary Figure 4).
5-ethynyl-2′-deoxyuridine EdU (Invitrogen) can efficiently label cycling cells in hydra as seen with a short pulse of three hours (supplementary material Fig. S1A), during which EdU was rapidly incorporated in dividing cells in the body column.
