The optimal concentration of each protein antigen was determined separately by visual examination of the signal versus the noise by using a positive serum sample pool and two negative serum samples (one sample was from a patient with echinococcosis and the other was a negative serum sample pool). The optimum concentrations of antigens were determined to be as follows: rGP50, 0.1 ng/mm; rT24H, 2.5 ng/mm; sTsRS1, 2 ng/mm; sTs18var1, 0.45 ng/mm; sTsRS2var1, 0.5 ng/mm; sTs14, 3 ng/mm; rES38, 1.25 ng/mm; and rES33, 2.5 ng/mm (Fig. ).
FIG. 1. MAPIA with cysticercosis and taeniasis antigens. Images of strips printed with eight individual antigens and developed after incubation with the cysticercosis-positive serum pool (lane 1), serum from a patient with echinococcosis (lane 2), and serum from (more ...)
Reading of the MAPIA strips was straightforward, and the level of agreement between the two readers was high. The kappa values for rGP50, rT24H, sTsRS1, sTs18var1, sTsRS2var1, sTs14, rES38, and rES33 were 0.98, 0.98, 0.96, 0.96, 0.91, 0.98, 0.92, and 0.98, respectively.
We evaluated the sensitivity of the individual antigens using sera from patients with clinically confirmed cases of cysticercosis. The rT24H antigen had the highest reactivity for all presentations of cysticercosis (Table ). The four 8-kDa antigens showed similar reactivities with the cysticercosis-positive sera. They were also the least recognized proteins among the larval-stage proteins by sera from all different categories of patients with neurocysticercosis. The positivity of the sera was the highest with sera from cases with two or more viable cysts and was the lowest with sera from cases with only calcified cysts. In sera from cases with two or more viable cysts, the use of more than one antigen did not improve the sensitivity of detection. Overall, the use of rT24H alone was sufficient. However, for the detection of cases with a single viable cyst, any combination of antigens resulted in a higher sensitivity compared to that achieved with the use of rT24H alone (sensitivities, 67.7% and 60%, respectively), and the increase in sensitivity was contributed by a combination of all four 8-kDa antigens. No specific reactivity patterns could be correlated to the different clinical presentations of neurocysticercosis (e.g., the presence of calcified or degenerating cysts).
Sensitivity of MAPIA with cyst-stage proteins and peptides for detecting neurocysticercosis
We evaluated the specificities of the individual antigens using a serum panel that consisted of sera from healthy individuals and sera from cases diagnosed with parasitic diseases other than cysticercosis and taeniasis. rGP50 had the greatest specificity; no sera from the specificity panel reacted with the antigen (Table ).
Specificity of MAPIA with cyst-stage proteins and peptides
To evaluate the ability of the recombinant adult worm antigens to detect taeniasis, we evaluated sera from cases with confirmed taeniasis and looked for reactivity with two tapeworm-specific proteins, rES38 and rES33. Of 162 specimens tested, 1 serum specimen did not react to either rES38 or rES33 (sensitivity of the taeniasis MAPIA, 99.4%). The specificity was evaluated by using the specificity panel; and the specificities of rES33 and rES38 were 93.9% and 94.5%, respectively; 17 serum samples from cases with schistosomiasis reacted with rES38; 19 serum samples reacted with rES33. Sera from six cases of confirmed Taenia saginata infection did not react with rES38 or rES33 (Table ). In this matter, use of a combination of Taenia antigens did not improve the sensitivity or the specificity. Although the sera from patients with cysticercosis and taeniasis were collected on the basis of the findings of cysts by a CT scan or MRI and the recovery of adult T. solium worms, respectively, these defined sera were not tested at the same time for both infections. By using the sera from cases with cysticercosis and two or more viable cysts, 81% of the subjects had antibody against rES33 and 83% had antibody against rES38. In subjects with active taeniasis, the reactivities to rGP50, rT24H, sTsRS1, sTs18var1, sTsRS2var1, and sTs14 were 88.9%, 93.2%, 35.2%, 34%, 42%, and 29.6%, respectively. All of these 53 cases, including 25 cases without neurological symptoms and negative neuroimaging findings, were seropositive for cysticercosis and taeniasis antigens.
Sensitivity and specificity of MAPIA with T. solium-derived adult tapeworm antigens