In a paper published recently in BMC Veterinary Research, Gertie Bokken and Frans van Knapen of Utrecht University in the Netherlands and Aldert Bergwerffs of Ghent University in Belgium describe the development of a novel, bead–based flow cytometric assay for the simultaneous determination of antibody responses against Toxoplasma gondii and Trichinella spiralis in pig serum.

The researchers explain that T. spiralis and T. gondii are well known zoonoses, which can pass from pigs to humans by consumption of raw or undercooked infected pork. In humans, most cases go undetected or lead only to mild disease but some cases can be very severe.

This high throughput screening assay could be an alternative for well known indirect tests like ELISA, according to the researchers.

One of the advantages of a bead-based assay over ELISA is the possibility to determine multiple specific antibody responses per single sample run facilitated by a series of antigens coupled to identifiable bead-levels. Furthermore, inclusion of a non-coupled bead-level in the same run facilitates the determination of, and correction for non-specific binding.

The performance of the bead-based assay was compared to one T. spiralis and three T. gondii ELISAs. For this purpose, sera from T. gondii and T. spiralis experimentally infected pigs were used.

With the experimental infection status as gold standard, the area under the curve, Youden Index, sensitivity and specificity were determined through receiver operator curve analysis. Marginal homogeneity and inter-rater agreement between bead-based assay and ELISAs were evaluated using McNemar’s Test and Cohen’s kappa, respectively.

Results indicated that the areas under the curve of the bead-based assay were 0.911 and 0.885 for T. gondii and T. spiralis, respectively, while that of the T. gondii ELISAs ranged between 0.837 and 0.930 and the T. spiralis ELISA was 0.879.

Bead-based T. gondii assay had a sensitivity of 86 per cent and specificity of 96 per cent, while the ELISAs ranged between 64 to 84 per cent and 93 to 99 per cent, respectively.

The bead-based T. spiralis assay had a sensitivity of 68 per cent and specificity of 100 per cent while the ELISA scored 72 per cent and 95 per cent, respectively.

Marginal homogeneity was found between the T. gondii bead-based test and one of the T. gondii ELISAs. Moreover, in this test combination and between T. spiralis bead-based assay and respective ELISA, an excellent inter–rater agreement was found. When results of samples before expected seroconversion were removed from evaluation, notably higher test specifications were found.

Bokken and co-authors concluded that this new bead-based test, which detects T. gondii and T. spiralis antibodies simultaneously within each sample, can replace two indirect tests for the determination of respective antibodies separately, while performing equally well or better.

Reference

Bokken G.C.A.M., A.A. Bergwerff and F. van Knapen. 2012. A novel bead-based assay to detect specific antibody responses against Toxoplasma gondii and Trichinella spiralis simultaneously in sera of experimentally infected swine. BMC Veterinary Research, 8:36. doi:10.1186/1746-6148-8-36

Further Reading

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You can view the full report (as a provisional PDF) by clicking here.