Abstract
BackgroundCurrently, the reference standard assay for the serodiagnosis of neurocysticercosis (NCC) is the lentil lectin-bound glycoproteins/enzyme-linked immunoelectrotransfer blot (LLGP-EITB). The main disadvantage of this technique is the complexity of obtaining and purifying the LLGP extract. This could be solved by replacement with highly specific recombinant antigens from Taenia solium. Based on previous studies, we selected and produced the recombinant Ts8B2 and T24H proteins and applied them to three diagnostic techniques: western blot (WB), enzyme-linked immunosorbent assay (ELISA) and the multiplex bead-based assay (MBA).MethodsThe Ts8B2 and T24H cDNA sequences were expressed in a prokaryotic system and the corresponding expression products purified; three recombinant proteins were further characterized: T24H-his, GST-T24H and GST-Ts8B2. The proteins on WB, ELISA and MBA were tested against 149 sera from patients with NCC confirmed by brain imaging, 40 sera from patients with other parasitic diseases, and 131 sera from US. individuals without evidence of neurocysticercosis (clinical/serological/brain imaging). The sensitivity and specificity of each antigen by WB were calculated by counting the number of true positive, false positive, true negative and false negative results. Using the receiver operating characteristic (ROC) curves, the cut-off values for the ELISA and MBA were established as well as the sensitivity and specificity of each assay.ResultsAll three antigens showed a high sensitivity on WB in active NCC cases with two or more viable cysts and low sensitivity for cases with single viable cyst or calcified lesions and inactive NCC. WB showed the highest specificity and sensitivity out of the three diagnostic techniques. The recombinant T24H-his was the best diagnostic reagent in WB (100% sensitivity, 99.4% specificity), exhibiting similar results to the LLGP-EITB, against the same panel of NCC sera. The GST-T24H antigen worked better than the others in ELISA and MBA protocols (88.3 and 96.1% sensitivity, respectively and 96.5% specificity).ConclusionsThe sensitivity and specificity that we obtained were similar to results from a previous study using a similar recombinant antigen (rT24H), suggesting that recombinant antigens may be good alternatives to crude extracts in a variety of diagnostic techniques. Furthermore, these antigens can be applied in the development of point-of-care tests which would be useful in NCC field studies.
Highlights
The reference standard assay for the serodiagnosis of neurocysticercosis (NCC) is the lentil lectin-bound glycoproteins/enzyme-linked immunoelectrotransfer blot (LLGP-enzyme-linked immunoelectrotransfer blot assay (EITB))
The objectives of this study were to obtain highly specific and sensitive recombinant antigens and to evaluate their application to various techniques for NCC serodiagnosis, in order to: (i) replace the use of T. solium crude extracts, and (ii) have different tests available according to the diagnostic needs; for instance, techniques to analyze a large number of samples, assays to discriminate the different NCC forms, tests to be developed using new materials or with innovative methodologies techniques easy to handle in endemic regions, etc
The recombinant proteins GST-T24H and T24H-his are good alternatives to the lentil lectin-bound purified glycoproteins (LLGP) antigen for the diagnosis of NCC, since the results were very similar for the recombinant molecules and the reference purified antigens in the same techniques, western blot (WB) or EITB
Summary
The reference standard assay for the serodiagnosis of neurocysticercosis (NCC) is the lentil lectin-bound glycoproteins/enzyme-linked immunoelectrotransfer blot (LLGP-EITB). The main disadvantage of this technique is the complexity of obtaining and purifying the LLGP extract. This could be solved by replacement with highly specific recombinant antigens from Taenia solium. The diagnosis must be based on a combination of the results obtained by imaging, serological tests and exposure history [7, 8]. The reference standard is lentil lectin-bound purified glycoproteins (LLGP) in an enzyme-linked immunoelectrotransfer blot assay (EITB) [7, 9]
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