Improving diagnosis and oral vaccination strategies against bovine tuberculosis

In this work peptide antigens [ESAT-6,p45 in water (1ml, 1mg/ml)] have been adsorbed onto 10mg inorganic substrates (hydroxyapatite (MHA P201;P120, CHA), polystyrene, calcium carbonate and glass microspheres) and in vitro release characteristics were determined. The aim of formulation was to enhance the interaction of peptides with antigen presenting cells and to achieve rapid peptide release from the carrier compartment system in a mildly acidic environment. Hydroxyapatite microparticle P201 has a greater surface area and thus has the largest peptide adsorption compared to the P120. CHA gave a further higher adsorption due to larger surface area than that available on microparticles. These particles were incorporated into the BOVIGAMTM assay to determine if they improve the sensitivity. After overnight incubation the blood plasma was removed and the amount of IFN-g in each plasma sample was estimated. CHA and MHA P201 gave a significantly higher immune response at low peptide concentration compared to the free peptide, thus indicating that these systems can be used to evaluate Tuberculosis (TB) amongst cattle using the BOVIGAMTM assay. Badgers are a source of TB and pass infection to cattle. At the moment vaccination against TB in badgers is via the parenteral route and requires a trained veterinary surgeon as well as catching the badgers. This process is expensive and time consuming; consequently an oral delivery system for delivery of BCG vaccines is easier and cheaper. The initial stage involved addition of various surfactants and suspending agents to disperse BCG and the second stage involved testing for BCG viability. Various copolymers of Eudragit were used as enteric coating systems to protect BCG against the acidic environment of the stomach (SGF, 0.1M HCl pH 1.2 at 37oC) while dissolving completely in the alkaline environment of the small intestine (SIF, IM PBS solution pH 7.4 at 37oC). Eudragit L100 dispersed in 2ml PBS solution and 0.9ml Tween 80 (0.1%w/v) gave the best results remaining intact in SGF loosing only approximately 10-15% of the initial weight and dissolving completely within 3 hours. BCG was incorporated within the matrix formulation adjusted to pH 7 at the initial formulation stage containing PBS solution and Tween 80. It gave viability of x106 cfu/ml at initial formulation stage, freezing and freeze-drying stages. After this stage the matrix was compressed at 4 tons for 3 mins and placed in SGF for 2 hours and then in SIF until dissolved. The BCG viability dropped to x106 cfu/ml. There is potential to develop it further for oral delivery of BCG vaccine.

The effect of stimulus location on the major components of the visual evoked response

The topographical distribution of the pattern reversal Visual Evoked Response (VER) was recorded from a localised montage of 20 electrodes over the visual cortex. The response was recorded after stimulation with a black and white checkerboard stimulus. The effect of field location on the major components was investigated in 11 subjects (age range (23-55). The major components of the half field response were; a negative around 75ms (N75) followed by a positivity around 80ms (P80), then a positivity around 100ms (P100) followed by another positivity at around 120ms (P120) and a negativity at approximately 145ms (N145). No effect of field size could be demonstrated on either the amplitude or latency of the late negativity, N145. No significant effect of field size or location was shown on the latency of the P100 response. A delay previously shown in the upper half field response was therefore not substantiated. In contrast the amplitude of the major positivity, P100 was significantly affected by the field size and location. The amplitude of both P100 and N145 were significantly reduced following upper field stimulation when compared with the lower field response. No significant amplitude difference between the upper and lower field responses was demonstrated using electroretinography, the amplitude may therefore be reduced as a result of the ventral position of the upper field representation on the visual cortex. The lateral half field VEP was compared with the distribution of the visual evoked magnetic response (VEMR). The distribution of the VEMR supported the proposal that the paradoxical lateralisation of the VEP half field response is the result of the source being directed ipsilaterally. The morphology of the VEP following octant and double octant stimulation suggests that the response is generated in the striate cortex, with a reversal in response distribution following stimulation of the upper vertical and horizontal meridia.