MSIPS4510 MilliporeMultiScreen_HTS IP Filter Plate, 0.45 µm, clear, sterile

The immune response induced by intradermal vaccination using a needle-less device was evaluated in conventional pigs in comparison with the more conventional intramuscular vaccination; to this purpose, vaccination against Aujeszky's Disease (AD) was used as a model of antiviral immunity. Two groups of pigs (n=10 each) were vaccinated 4weeks apart respectively by the intramuscular (IM group) and intradermal route (ID group; needle-less I.D.A.L.(R) vaccinator) with an AD modified live virus. Ten pigs injected with the vaccine adjuvant only were kept as sham-vaccinated controls (C group). On blood samples collected at 0, 2, 4, 5, 6 and 7weeks post-vaccination (PV) ADV-specific virus neutralizing (VN) antibodies, IFN-gamma secreting cells (SC), lymphocyte subsets and IFN-gamma gene expression in PBMC were evaluated. VN antibodies increased after the 1st vaccination and peaked after the 2nd vaccination in both vaccinated groups. Also IFN-gamma SC reached maximum levels in both groups after administration of the booster dose. Pigs in the control group remained negative for both parameters throughout the study. Flow cytometry showed persistently higher levels of CD3-CD8alpha+ Natural Killer cells in both vaccinated pigs. The ID group showed an earlier and regulated activation characterized by an increase of cytotoxic CD8beta+ T lymphocytes and CD25+ cells after the boosting dose. No statistically significant differences between treated and control groups were detected for memory CD4+CD8alpha+(low) T cells. Upregulation of IFN-gamma gene expression in PBMC was detected in ID and IM pigs after both vaccine administrations, although at a different extent. Overall, the results showed that the intradermal vaccine delivery by a needle-less device can prime a strong humoral and cellular immune response comparable to that obtained by the intramuscular vaccination.

Assays detecting antigen (Ag)-specific T-cell responses in immune-mediated processes are increasingly employed to understand disease pathogenesis and immune staging. The quantity and quality of starting peripheral blood mononuclear cell (PBMC) preparations are important factors in the performance of such assays. We therefore compared final PBMC yield and function by modifying parameters at the blood drawing, storage and processing steps. While drawing blood in vacuum-driven tubes or syringes and separating PBMCs on density gradients using standard or membrane (Leucosep) tubesmade no difference, storing tubes for 18 h without any agitation led to PBMC preparations contaminated with granulocytes and decreased interferon (IFN)-gamma enzyme-linked immunospot (ELISpot) responses. Even agitated blood showed a trend towards reduced ELISpot responses and increased human leukocyte Ag (HLA) multimer readouts when stored for 18 h compared to 3 h. These changes were reduced by diluting blood prior to storage. Washing PBMCs with media containing 10% human serum increased PBMC yields by 40.5%, without affecting ELISpot responses and multimer counts. However, washes with > 10% human serum decreased multimer counts, with no additional improvement in PBMC yields. These findings may be relevant for optimizing and harmonizing PBMC processing procedures for T-cell assays.

This study investigated the efficacy of a one-dose porcine circovirus 2 (PCV2) subunit vaccine based on the PCV2 Cap protein expressed in a baculovirus system on two different farms at which a history of porcine circovirus-associated disease (PCVD) was present. Morbidity, mortality, average daily weight gain, carcass weight, PCV2 load in serum and vaccine immunogenicity were assessed. Serology to porcine reproductive and respiratory syndrome virus (PRRSV) and Mycoplasma hyopneumoniae was performed. A double-blind, randomised, and controlled field trial was performed distributing 818 piglets between two treatment groups. At inclusion (weaning at 21±3 days of age), 408 animals (group B) received a 2-mL intramuscular dose of Porcilis PCV(®) (vaccinated group). Controls (group A, 410 pigs) received 2mL of the adjuvant Diluvac Forte(®) intramuscularly. Weights were recorded at inclusion and at 12 and 26 weeks of age, and the average daily weight gain (ADWG) was calculated. The carcass weights of the pigs from farm 2 were recorded at slaughter (274 days old). All dead animals (died or culled) underwent autopsy to classify them as PMWS-affected or not. At each farm, blood samples were taken from 22 pigs/group for serologic studies. A beneficial effect was found after vaccination with a single dose of a PCV2 Cap vaccine against PCVD. The vaccination reduced the mortality rate and morbidity, reduced PCV2 viremia and viral load, improved productive performances (e.g. ADWG: +70g/day between 12 and 26 weeks of age when viremia and the specific disease occurred) as well as carcass weight at slaughter age (+4.5kg). These effects were associated with virologic and clinical protection from the immunogenicity of the vaccine measured as activation of both a humoral and a cellular immune response.