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Abstract
An effective contagious caprine pleuropneumonia (CCPP) vaccine is essential for the increased production of healthy goats in a cost-effective manner and the prevention of animal-to-animal transmission for both domestic animals and wildlife. Quality control of this vaccine ensures that a reliable supply of pure, safe, and potent batches is obtained. As part of this control, in vitro quantification of Mycoplasma capricolum subsp. capripneumoniae (Mccp) protein in the final vaccines is required before the CCPP vaccine undergoes batch release and certification. The current method used for quantification is based on the measurement of total protein using the bicinchonic acid (BCA) test. This method quantifies the total amount of protein in the vaccine including contaminant protein from media, which can lead to overestimation of the quantity of Mccp protein, resulting in reduced vaccine immunogenicity. An immuno-capture ELISA (ICE) was developed for specific detection and quantification of the Mccp antigen in the CCPP vaccine. As the ICE detects and measures the amount of antigen between two layers of antibodies, capture and detecting antibodies are required. Mouse monoclonal antibodies (mAbs) that detect the Mccp antigen were produced and characterized. One of these mAbs, Mccp-25, was used to develop the ICE as an unlabelled capture antibody and horseradish peroxidase conjugated detecting antibody. The ICE was standardized and evaluated using an internal reference sample, experimental CCPP vaccines and commercial CCPP vaccines. A comparison between the polymerase chain reaction (PCR) and ICE showed good correlation between the two assays. Also, an in vitro ICE method correlated well with an in vivo sero-conversion in goats that were vaccinated with selected test vaccines. The sensitivity of the ICE was estimated at 30 ng/ml.
Highlights
Contagious caprine pleuropneumonia (CCPP) is an infectious disease in goats caused by Mycoplasma capricolum subspecies capripneumoniae (Mccp)
For both forms, quantification of the Mccp antigen is used for vaccine batch potency evaluation. e optimum dose of antigen was established to be 0.15 mg of Mccp protein and 3 mg of saponin. e current assay used for Mccp antigen quantification in the CCPP vaccine is the bicinchonic acid (BCA) protein assay. is assay is used for quantitation of the total protein in a sample protein test and is not specific to the Mccp antigen
Culture and Molecular Characterisation of Mycoplasma capricolum Subspecies Capripneumoniae. e growth of Mccp was observed by the presence of filaments and the turbidity of the media. e monitoring of the pH of the Mccp culture media showed a slight acidity compared to the pH of the normal media (Figure 1). e noninoculated Pleuropneumonia-Like Organisms (PPLO) broth
Summary
Contagious caprine pleuropneumonia (CCPP) is an infectious disease in goats caused by Mycoplasma capricolum subspecies capripneumoniae (Mccp). E CCPP vaccine is an inactivated preparation of F38 mycoplasma containing saponin as an inactivating agent and adjuvant [13]. E liquid form makes this formulation less stable than the freeze-dried form For both forms, quantification of the Mccp antigen is used for vaccine batch potency evaluation. E current assay used for Mccp antigen quantification in the CCPP vaccine is the bicinchonic acid (BCA) protein assay. The development of an ICE assay using a mAb as a capture and detecting antibody for the detection and quantification of the Mccp protein in the CCPP vaccine is reported
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