Anionic microgels have been widely used in drug delivery and surface modification. Their ability to complex with oppositely charged molecules has drawn increasing attention. Many imaging techniques have successfully been used to study microgels including AFM and confocal microscopy, among others. Scanning electron microscopy is particularly appropriate because of its combination of resolution, depth of field, and microanalytical abilities. Characterizing microgels by SEM has, however, been limited because of drying artifacts. Here, we prepare frozen-hydrated samples using high-pressure freezing. We use this approach to study the complexation of cationic antimicrobials within anionic microgels where imaging and X-ray microanalysis (EDS) provide direct evidence for water and counterion release during complexation.<br/><br/>Our experiments use a cationic antimicrobial peptide L5 (+6 charge, 2274 g/mol) and lightly crosslinked poly(acrylic acid) microgels. L5-complexed microgels form a core-shell structure. Significantly different X-ray intensity profiles in the core and the shell indicate differences in local composition. The carbon and oxygen X-ray intensities indicate that the core is highly hydrated while the shell is highly dehydrated. Nitrogen is unique to L5, and a spike in the nitrogen X-ray intensity indicates that the peptide is localized preferentially in the shell. Also, a depletion of sodium is observed exclusively in the shell. These findings indicate that peptide-microgel complexation drives the local release of both water and sodium counter ions. By studying specimens frozen after various complexation times we find that longer complexation results in a thicker and more dehydrated shell with sufficient mechanical integrity to resist deformation during complete sublimation. The rate of shell thickening decreases such that we observe a maximum thickness of about ~4 µm. The overall peptide-complexed microgel diameter is ~50 µm, however, and the much larger core remains hydrated and with no significant changes.

Cryo-SEM imaging and analysis of peptide-complexed microgels

256th National Meeting (2018)

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