Method for quantifying the Pasteurella multocida antigen adsorbed on aluminum hydroxide adjuvant in swine atrophic rhinitis vaccine.

Swine atrophic rhinitis is a disease caused by Pasteurella multocida and Bordetella bronchiseptica that affects pigs. Inactivated vaccines containing the toxins produced by Pasteurella multocida and Bordetella bronchiseptica have been widely used for the prevention of swine atrophic rhinitis. The efficacy of a vaccine is correlated with the amount of antigen present; however, the protective toxin of P. multocida bound to aluminum hydroxide, which is used as an adjuvant, can hinder the monitoring of the antigen concentration in the vaccine. This study assessed the applicability of a dot immunoassay as an antigen quantification method using monoclonal antibodies. This quantification method was able to detect the antigen with high specificity and sensitivity even when the antigen was bound to the adjuvant, and its application to vaccine products revealed a correlation between the amount of antigen present in the vaccine and the neutralizing antibody titers induced in pigs. The antigen quantification method presented in this study is a simple and sensitive assay capable of quantifying the amount of antigen present in a vaccine that can be used as an alternative quality control measure.

Induction of antitoxic antibody and preventive effect against porcine edema disease by the pentameric Stx2eB subunit vaccine.

Porcine edema disease (ED) is an enterotoxaemia that frequently occurs in 4-12 week-old piglets and results in high mortality. ED is caused by Shiga toxin 2e (Stx2e), produced by host-adapted Shiga toxin-producing Escherichia coli (STEC) strains. We constructed a recombinant protein in which the B subunit of Stx2e (Stx2eB) was linked to Cartilage Oligomeric Matrix Protein (COMP)'s pentameric domain to enhance antigenicity to induce neutralizing antibodies against Stx2e. We evaluated the efficacy of this antigen as a vaccine on the farm where ED had occurred. The suckling piglets were divided into two groups. The pigs in the vaccinated group were intramuscularly immunized with the vaccine containing 30 µg/head of Stx2eB-COMP at 1 and 4 weeks of age. The control pigs were injected with saline instead of the vaccine. The neutralizing antibody titer to Stx2e, mortality, clinical score, and body weight was evaluated up to 11 weeks after the first vaccination. In the vaccinated group, the Stx2e neutralizing antibody was detected 3 weeks after the first vaccination, its titer increased during the following weeks. The antibody was not detected in the control group during the test period. The STEC gene was detected in both groups during the test period, but a typical ED was observed only in control pigs; the mortality and clinical score were significantly lower in the vaccinated group than in the control group. These data indicate that the pentameric B subunit vaccine is effective for preventing ED and offers a promising tool for pig health control.

Effects of various methoxyflavones on vincristine uptake and multidrug resistance to vincristine in P-gp-overexpressing K562/ADM cells.

PURPOSE: Some methoxyflavones (MFs) are known to inhibit the function of P-glycoprotein. The aim of this study is to characterize the reversal of multidrug resistance (MDR) by MFs. METHODS: The effects of 19 MFs, including 3,5,6,7,8,3',4'-heptamethoxyflavone, nobiletin, and tangeretin, and flavone on the uptake of [3H]vincristine into an adriamycin-resistant variant of human chronic myelogenous leukemia (K562/ADM) cells were investigated. Potentiation of vincristine-induced growth inhibition by these MFs was also tested in K562/ADM cells by means of WST-1 [2-(4-iodophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium] assay. RESULTS: All MFs (20 microM) tested increased the uptake of [3H]vincristine. 3,5,6,7,8,3',4'-heptamethoxyflavone, nobiletin, tangeretin, quercetagetin and quercetin pentamethylether showed especially potent effects. The increase in the uptake of [3H]vincristine was proportional to the number of methoxyl moieties. While substitution with a methoxyl moiety at the C3 position was the most influential, methoxyl substitution at both the C3' and C5' positions resulted in a decrease in the potentiation of uptake. Furthermore, there was a significant correlation between the potencies for increasing [3H]vincristine uptake and for growth inhibition assessed by WST-1 assay. CONCLUSIONS: MFs increased the uptake of [3H]vincristine into MDR cells and exhibited MDR-reversing effects. Their potencies were influenced by the number and positions of the methoxyl moieties.