Recombinant Protein- and Synthetic Peptide-Based Immunoblot Test for Diagnosis of Neurocysticercosis

Abstract

One of the most well-characterized tests for diagnosing neurocysticercosis (NCC) is the enzyme-linked immunoelectrotransfer blot (EITB) assay developed at the CDC, which uses lentil lectin-bound glycoproteins (LLGP) extracted from Taenia solium cysticerci. Although the test is very reliable, the purification process for the LLGP antigens has been difficult to transfer to other laboratories because of the need for expensive equipment and technical expertise. To develop a simpler assay, we previously purified and cloned the diagnostic glycoproteins in the LLGP fraction. In this study, we evaluated three representative recombinant or synthetic antigens from the LLGP fraction, individually and in different combinations, using an immunoblot assay (recombinant EITB). Using a panel of 249 confirmed NCC-positive and 401 negative blood serum samples, the sensitivity of the recombinant EITB assay was determined to be 99% and the specificity was 99% for diagnosing NCC. We also tested a panel of 239 confirmed NCC-positive serum samples in Lima, Peru, and found similar results. Overall, our data show that the performance characteristics of the recombinant EITB assay are comparable to those of the LLGP-EITB assay. This new recombinant-and synthetic antigen-based assay is sustainable and can be easily transferred to other laboratories in the United States and throughout the world.T he diagnosis of neurocysticercosis (NCC), a condition caused by the larvae Taenia solium, is reliably established using results from both imaging and serological tests. The guidelines proposed for making a diagnosis of presumptive NCC (1, 2) define cases as definitive, probable, or possible based on computed tomography (CT) or magnetic resonance imaging (MRI) findings, exposure risk, and serological results. These guidelines specifically define the lentil lectin-bound glycoprotein enzyme-linked immunoelectrotransfer blot assay (LLGP-EITB) as the serological reference standard for diagnosing NCC (3).The LLGP-EITB assay is an immunoblot method that detects antibodies to one or more of seven lentil lectin-bound glycoproteins, which are present in the soluble fraction of an extract of T. solium cysts. In patients with multiple enhancing intracranial lesions, the LLGP-EITB assay is 100% specific and 95% sensitive using blood serum or cerebrospinal fluid (CSF) samples (3-6). The original study that described and evaluated the LLGP-EITB assay was performed using serum samples from biopsy-proven cases of NCC, most with multiple lesions, as detected by skeletal radiographs, and reported a sensitivity of 98% and a specificity of 100% (3). Continued monitoring of the test performance compared with clinical findings using newer imaging techniques, such as CT and MRI, revealed that the sensitivity of the assay was lower, between 50 to 80%, in cases with a single lesion or calcified cysts (6-10). The specificity of the LLGP-EITB assay has been remarkable at essentially 100%, with only rare anecdotal reports of falsepositive results (11)(12)(13).Although the LLGP-...

Low sensitivity and frequent cross‐reactions in commercially available antibody detection ELISA assays for Taenia solium cysticercosis

“…While we selected two ELISA assays easily available in the market, we may have missed other less known kits with better performance. There are reports in the literature of recently identified antigens that perform well in exploratory testing and have improved specificity including recombinant and synthetic molecules [25][26][27][28][29][30][31][32][33][34][35][36] but to the best of our knowledge, ELISAs using these antigens are not yet commercially available (Akira Ito and Sukwan Handali, personal communication, 2017). Theoretically, a highly specific antigen could be used in higher concentrations and thus potentially also improve assay sensitivity and overall assay performance.…”

Association between Taenia solium infection and HIV/AIDS in northern Tanzania: a matched cross sectional-study

“…LLGP-EITB is an enzyme-linked immunoelectrotransfer blot that detects CC-specific Ab to any of seven glycoproteins [41, 42]. The rT24H-test is an immunoblot method that detects CC-specific Ab to a T. solium recombinant Ag [43]. The Ag-ELISA that detects Taenia -Ag in serum is a monoclonal Ab (B158/B60 Ab) based capture ELISA and was performed following the modified protocol described by Dorny et al [44].…”

Comparison of T24H-his, GST-T24H and GST-Ts8B2 recombinant antigens in western blot, ELISA and multiplex bead-based assay for diagnosis of neurocysticercosis

“…The recombinant GP50 and T24H [14, 15], derived from the 50 kDa and 24–42 kDa bands from the LLGP, have been tested in techniques such as multi antigen printing immunoassay (MAPIA), ELISA, western blot, QuickELISA and rapid Lateral Flow assay [15–18], with equally high specificity and sensitivity comparable to the reference standard [19]. …”