Silicon nitride cantilevers

The morphology
and size of the peptide-pNIPAm SAPEs were visualized by transmission
electron microscopy (TEM). Briefly, samples (0.1 mg/mL in HEPES 10
mM) were soaked onto prewarmed carbon-coated copper grids for 2 min,
and excess liquid was removed by a filter paper. The grids were subjected
to negative staining with 2% uranyl acetate (Merck, Germany) for 45
s and dried for 10 min at room temperature before acquisition of TEM
images (Tecnai 10, Philips, The Netherlands). Atomic force microscopy
(AFM) was applied to visualize the morphology of the peptide-based
vaccines. Briefly, 40 μL of a 0.02% (w/v) polylysine solution
in water was placed on a freshly cleaved mica substrate and, after
5 min incubation, washed with filtered Milli-Q water followed by drying
under N₂ flow. Next, 30 μL of the sample was loaded
on the treated mica and incubated for 1 min, followed by washing three
times with 20 μL of filtrated Milli-Q water to discharge nonattached
particles and salts. Finally, the prepared sample was dried by an
N₂ flow. ScanAsyst image mode was used at room temperature
with a Digital Instruments NanoScope V, equipped with silicon nitride
cantilevers (Veeco, NY, USA). Images were analyzed by NanoScope Analysis
1.40 software. Several images were taken at different days of individually
prepared samples.