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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.

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.

Enzyme linked immunosorbent assays (ELISAs) have been developed for the detection of antibodies against contagious caprine pleuropneumonia (CCPP), the causative agent of which is Mycoplasma capricolum subsp. Capripneumoniae (Mccp). The currently available commercial CCPP competitive ELISA (CCPP cELISA) kit produced and supplied by IDEXX Company (Westbrook, Maine, United States) is relatively expensive for most African laboratories. To address this issue and provide a variety of choices, a sensitive and specific blocking-ELISA (b-ELISA) test to detect antibodies against CCPP was developed. We describe the newly developed CCPP blocking-ELISA based on the blocking of an epitope of a monoclonal antibody (Mccp-25) by a positive serum sample against the Mccp protein coated on a plate. The Percentage Inhibition (PI) cut-off value for the CCPP b-ELISA was set at 50 using 466 CCPP negative and 84 CCPP positive small ruminant sera. Of the negative sera, 307 were obtained from the Botswana National Veterinary Laboratory (BNVL) and 159 from the Friedrich-Loeffler-Institute (FLI) Germany. The 84 positive sera samples came from experimentally vaccinated goats at the AU-PANVAC facility in Debre-Zeit, Ethiopia. The relative diagnostic sensitivity and specificity of the CCPP b-ELISA was 93% and 88%, respectively. This test result indicated good correlation with that of the commercial CCPP cELISA by IDEXX Company (Westbrook, Maine, United States) with a Cohen’s κ agreement of κ agreement of 0.85. The newly developed CCPP b-ELISA will be useful in the detection of antibodies for the diagnosis CCPP and for sero-surveillance during vaccination campaigns.