The cells were filtered through a 70-μm cell strainer to obtain single cell suspension before analyzed with a LSRII flow cytometer (BD Biosciences, Franklin Lakes, NJ, USA).
Before analyzing with SEM, the surface of algal cells was coated with a gold layer by employing Sputter Coater (Hitachi E-1010, Japan).
Fixed cells were then stained with the cyclin B1 antibody, as described earlier, except that Hoechst was used, and the cells were kept in PBS before analyzing with the HCS system.
Cells were washed twice in assay buffer before analyzed with flow cytometer.
They were then vacuum-dried and digested with 8 µL trypsin solution before being analyzed with a 4700 Proteomics Analyzer.
Blood samples from the day of VAP diagnosis and 1 and 3 days before were analyzed with a multiplex array for markers of inflammation, coagulation, and apoptosis.
Cells were then washed, centrifuged and fixed with 500 µl of 1% formaldehyde before being analyzed with a FACS Calibur cytometer (Becton Dickinson, Le Pont de Claix, France).
Dehydration was achieved using an ethanol scale and samples were gold coated before being analyzed with a Zeiss EVO 50 environmental scanning electron microscope (ESEM).
The sample was then incubated in the dark at room temperature for 30 min before being analyzed with a BD FACScan™ flow cytometer (BD Biosciences, San Jose, CA) using CellQuest Software (BD Biosciences, San Jose, CA).
Dried samples from the degradation study and the swelling ratio study (24 h in PBS before being lyophilized) were analyzed with a Nicolet FTIR microscope.
