An enzyme-linked immunosorbent assay (ELISA) employing protein AG (AG-ELISA) as a conjugate was developed to detect anti-Toxoplasma gondii antibodies (Ab) in experimentally infected pigs and naturally infected pigs, goats, dogs, and cats. The results indicate that AG-ELISA can be a useful method for serological diagnosis of T. gondii infection in these four species of animals.Infections with Toxoplasma gondii are prevalent in human beings and animals worldwide (1). Numerous studies of T. gondii infections in pigs and other animals have demonstrated that enzyme-linked immunosorbent assay (ELISA) is the most sensitive method for diagnosing T. gondii infection (2,4,8). However, ELISA needs species-specific secondary antibodies for testing each species of animals. This requirement increases cost and makes manipulation more complicated when samples from different animal species are examined. Since the chimeric protein AG can strongly bind to the IgG of many mammalian species, it has been a powerful tool in immunological studies (5, 6). So far, there has been no report on the utility of chimeric AG in the immunological diagnosis of T. gondii infection. In this study, chimeric protein AG was employed to develop a simple protein AG enzyme-linked immunosorbent assay (AG-ELISA) to detect T. gondii antibodies from different animal species.For establishing the AG-ELISA method, recombinant MIC3, which was highly expressed in Escherichia coli by our laboratory as previously described (9), was used as the coating antigen. Flat-bottomed 96-well polystyrene micro-titration plates were coated with 0.1 ml of the antigens (2.5 mg/liter) diluted in 0.05 M carbonate buffer (pH 9.6) by incubation overnight at 4°C and were blocked with carbonate buffer-1% ovalbumin for 1 h at 37°C. The control and test sera were diluted 1:160 in phosphate-buffered saline-Tween (PBST) containing 0.1% ovalbumin, added to the microtiter plate at 0.1 ml per well, and incubated for 1 h at 37°C. Next, peroxidase-labeled chimeric protein AG (1:4,000; Pierce, Rockford, IL) was added at 0.1 ml per well and incubated for 30 min at 37°C. Peroxidase activity was revealed by adding 0.1 ml of tetramethylbenzidine (TMB) solution (100 mg TMB/liter of phosphate citrate buffer [pH 6.0] and 200 ul of H 2 0 2 ) for 10 min at room temperature. The reaction was stopped by adding 0.05 ml of 0.25% hydrofluoric acid (HF), and the optical density (OD) was read at 630 nm in an ELISA microplate reader. A serum sample was considered to be positive when the OD of the sample/the OD of the negative control was Ն2.3.Twelve mixed-breed pigs between 6 and 8 weeks old were purchased from one pig farm and randomly allocated to separate stables. The animals were acclimatized for 7 days before use. All pigs tested negative for the presence of T. gondii antibodies by AG-ELISA (OD Ͻ 0.16) and modified agglutination test (MAT) (titer Ͻ 1:16). At day 0, eight pigs were inoculated with 2 ϫ 10 4 of viable tachyzoites of the RH strain by the subcutaneous route; the other four pigs were used as con...
Toxoplasma gondii is a protozoan parasite that causes severe diseases in mammals, including humans, around the world. In China, pork is the main meat source; accordingly, T. gondii in pigs is considered an important source for human toxoplasmosis. Understanding the epidemiology of toxoplasmosis in pig farms is thus important for control of the disease in humans. The purpose of the present study was to investigate the epizootiology of T. gondii infections in pig farms in central China by assessing the seroprevalence and risk factors of this disease. In the present study, 3,558 sera samples were collected from pigs in 37 large-scale pig farms in this region and tested by AG-ELISA. The total seroprevalence was 24.5%, with the greatest prevalence in breeding pigs. The risk factors for toxoplasmosis suggest that high frequency of the contact of pigs with cats (P ≤ 0.01; IC 95%), high density of pig breeding (P ≤ 0.01; IC 95%), the presence of mosquitoes and flies (P ≤ 0.01; IC 95%), semi-patency pens (P ≤ 0.05; IC 95%), and low frequency of scavenging (P ≤ 0.01; IC 95%) were all associated with seroprevalence. In addition, the use of sulfonamides (P ≤ 0.01; IC 95%) significantly decreased seroprevalence. This is the first report of anti- T. gondii antibodies in pigs on large-scale pig farms in central China. The findings will provide useful information for designing control strategies of toxoplasmasis in pig farms.
The pharmaceutical tablet manufacturing process (PTMP) via wet granulation holds a critical position in pharmaceutical industry. The interest in integrating mechanistic process modeling into the pharmaceutical development has been increased because simulation model is a prerequisite for process design, analysis, control, and optimization. So the simulation modeling for PTMP via wet granulation is very necessary and significant. This study aims at proposing a simulation modeling framework for PTMP via spray fluidized bed granulation (SFBG), which is one of the most widely used wet granulation techniques in pharmaceutical industry. For SFBG, a simulation model that simultaneously involves the influences of operating variables and material attributes on average particle size (APS) is firstly developed, and then a drying model to determine the particle moisture content is introduced to be coupled with the established model predicting APS. For PTMP, considering the important effect of porosity on tablet qualities, a model describing the changes in tablet porosity is developed based on a promoted form of the Heckel equation, and then several recognized models that are all related to porosity are introduced or constructed to calculate important tablet quality indexes. The feasibility and effectiveness of the developed simulation models are validated by performing a computational experimental study to explore the scientific understanding of process and process quality control.
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