Construction and physiochemical characterisation of a multi-composite, potential oral vaccine delivery system (VDS)

An increasing human population requires a secure food supply and a cost effective, oral vaccine delivery system for livestock would help facilitate this end. Recombinant antigen adsorbed onto silica beads and coated with myristic acid, was released (∼15% (w/v)) over 24 h at pH 8.8. At pH 2, the myristic acid acted as an enteric coating, protecting the antigen from a variety of proteases. The antigen adsorbed onto silica particles, coated in myristic acid had a conserved secondary structure (measured by circular dichroism (CD) spectroscopy) following its pH-triggered release. Small angle neutron scattering (SANS) was used to measure the thickness of the adsorbed antigen, finding that its adsorbed conformation was slightly greater than its solution radius of gyration, i.e. 120–160 Å. The addition of myristic acid led to a further increase in particle size, with scattering data consistent with an acid thickness slightly greater than a monolayer of fully extended alkyl chains and a degree of hydration of around 50%. Whilst adsorbed onto the silica and coated in myristic acid, the protein was stable over 14 days at 42 °C, indicating a reduced need for cold chain storage. These data indicate that further investigation is warranted into the development of this technology

Peptide-mass fingerprinting as a tool for the rapid identification and mapping of cellular proteins

For more than 25 years protein identification has largely depended on automated Edman chemistry (Hewick et al., 1981) or western blotting with an appropriate monoclonal antibody. Several limitations, however, have never been overcome. The Edman procedure is inherently slow (generally one or two peptide or protein samples per day) and does not allow direct identification of many post-translational modifications. In addition, current detection limits are in the low-picomole to upper-femtomole range (Totty et al., 1992). Protein identification by western blotting can be extremely rapid, but requires the ready availability of an extensive library of suitable antibody probes. Large-format 2D-electrophoresis systems now make it possible to resolve several thousand cellular proteins from whole-cell lysates in the low- to upper-femtomole concentration range (Patton et al., 1990), presenting significant analytical challenges. The recent introduction of matrix-assisted laser-desorption (MALD) time-of-flight mass spectrometers (Karas and Hillenkamp, 1988) has led to the rapid analysis (at high sensitivity) of peptide mixtures. New strategies have been developed using a combination of protease digestion, MALD mass spectrometry and searching of peptide-mass databases that promise rapid acceleration in the identification of proteins (Henzel et al., 1993; Pappin et al., 1993; Mann et al., 1993; James et al., 1993; Yates et al., 1993)