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The ratio of hydroxyapatite (HA) nanoparticles (NP) to trehalose in composite microparticle (MP) vaccine vehicles by determining inter-nanoparticle space potentially influences antigen release. Mercury porosimetry and gas adsorption analysis have been used quantify this space. Larger pores are present in MPs spray dried solely from nanoparticle gel compared with MPs spray dried from nanoparticle colloid which have less inter-nanoparticle volume. This is attributed to tighter nanoparticle packing caused by citrate modification of their surface charge. The pore size distributions (PSD) for MP where the trehalose has been eliminated by combustion generally broaden and shifts to higher values with increasing initial trehalose content. Modal pore size, for gel derived MPs is comparable to modal NP width below 30% initial trehalose content and approximates to modal NP length (~50 nm) at 60% initial trehalose content. For colloidally derived MPs this never exceeds the modal NP width. Pore-sizes are comparable, to surface inter-nanoparticle spacings observed by SEM.

Microparticles (MP) spray dried from hydroxyapatite (HA) nanoparticle (NP) sugar suspensions are currently under development as a prolonged release vaccine vehicle. Those with a significant sugar component cannot be sectioned by ultramicrotomy as resins are excluded by the sugar. Focused ion beam (FIB) milling is the only method to prepare thin sections that enables the inspection of the MPs ultrastructure by transmission electron microscopy (TEM). Several methods have been explored and we have found it is simplest to encapsulate MPs in silver dag, sandwiched between gold foils for FIB-milling to enable multiple MPs to be sectioned simultaneously. Spray dried MPs containing 80% sugar have an inter-nanoparticle separation that is comparable with NP size (~50 nm). MPs spray dried with 50% sugar or no sugar are more tightly packed. Nano-porosity in the order of 10 nm exists between NPs. MPs spray dried in the absence of sugar and sectioned by ultramicrotomy or by FIB-milling have comparable nanoscale morphologies. Selected area electron diffraction (SAED) demonstrates that the HA remains (substantially) crystalline following FIB-milling.