Comparison of enzyme-linked immunosorbent assay and indirect fluorescent antibody test for the detection of IgG antibodies to sarcocystis muris

Abstract

Sarcocystis muris cystozoites were separated from host tissue after mechanical mincing and homogenization by discontinuous density gradient centrifugation using Percoll®. The isolated antigen, consisting of more than 95% cystozoites, was used in an enzyme-linked immunosorbent assay (ELISA) and an indirect fluorescent antibody test (IFAT). In the ELISA, the antigen was used at a very low protein concentration of 0.44 μg/ml. Good reproducibility of test results was achieved with different conjugates and different reference sera when the measured optical densities were converted into an index related to a standard. From the examination of 500 sera from non-infected mice, an index of 0.300 at a serum dilution of 1:10 was determined as the threshold for a positive reaction in the ELISA. Specific antibodies were first detected between 18 and 49 days after experimental infection (dpi) with S. muris. The ELISA as well as the IFAT was highly sensitive from about 50 dpi onwards and detected antibodies up to the end of the examination period (182 dpi). However, the IFAT with unfixed antigen appeared more sensitive than the ELISA for the period of infection before 50 dpi and detected specific antibodies between 11 and 25 dpi starting with fluorescence at the apical pole of the cystozoite and resulting in bright fluorescence of the whole cystozoite from 32 or 35 dpi onwards. Both serotests showed only slight crossreactions with high titred Toxoplasma gondii sera at a serum dilution of 1:10. The activity of the antigen lasted for at least 13 months in the ELISA and for at least eight months in the IFAT.