title: Immunodiagnosis and molecular validation of Toxoplasma gondii-recombinant dense granular (GRA) 7 protein for the detection of toxoplasmosis in patients with cancer.
abstract: Serological assays for the diagnosis of toxoplasmosis mostly rely on the tachyzoite specific antigens of Toxoplasma gondii, which are difficult to produce by conventional methods. The aim of this study was to clone and express of GRA7 protein of T. gondii and evaluate its potential for immunodiagnosis of toxoplasmosis in cancer patients. As well as validate the results using a new molecular assay, LAMP technique. The GRA7 gene was successfully cloned, expressed and purified by affinity chromatography and the production was evaluated by SDS PAGE, dot blot and western blot analyses. The rGRA7 was used for developing an ELISA based on the rGRA7 using sera from patients with toxoplasmosis and healthy controls. Furthermore, 50 serum samples from leukemic children infected with toxoplasmosis and 50 seronegative controls were included to evaluate the sensitivity and specificity of rGRA7 based ELISA. Finally, the LAMP technique was used to assess the accuracy and validity of the results obtained by rGRA7 based ELISA. The consistency of the results of two tests was determined by using the Kappa coefficient of agreement. The rGRA7 showed higher and optimum immunoreactivity with 1:100 dilution of serum from Toxoplasma infected patients. The sensitivity and specificity of test were calculated as 92 and 94%, respectively. According to the Kappa coefficient of agreement, there was a significant conformance between the results obtained by ELISA based on the rGRA7 and the results of LAMP technique (≈96%, P<0.001). Findings of the present study showed that rGRA7 can be used as a potential immunogenic antigen for developing immunodiagnostic tools for immunodiagnosis of toxoplasmosis in patients including patients with cancer.
authors.affiliation: Infectious Diseases and Tropical Medicine Research Center, Shahid Beheshti University of Medial Sciences, Tehran, Iran; Department of Parasitology and Mycology, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
authors.forename: Zahra
authors.initials: Z
authors.lastname: Arab-Mazar
authors.affiliation: Department of Medical Parasitology and Mycology, Faculty of Medicine, Lorestan University of Medical Sciences, Khorramabad, Iran; Razi Herbal Medicines Research Center, Lorestan University of Medical Science, Khorramabad, Iran. Electronic address: Fallahi.Sh@lums.ac.ir.
authors.forename: Shirzad
authors.initials: S
authors.lastname: Fallahi
authors.affiliation: Department of Biotechnology, Faculty of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
authors.forename: Ameneh
authors.initials: A
authors.lastname: Koochaki
authors.affiliation: Department of Parasitology and Mycology, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
authors.forename: Ali
authors.initials: A
authors.lastname: Haghighi
authors.affiliation: Department of Parasitology and Mycology, Shahid Beheshti University of Medical Sciences, Tehran, Iran. Electronic address: Seyyedtabaei@sbmu.ac.ir.
authors.forename: Seyyed Javad
authors.initials: SJ
authors.lastname: Seyyed Tabaei
mesh_terms: D000953:Antigens, Protozoan / Q000032:analysis* / Q000097:blood / Q000235:genetics* / Q000302:isolation & purification; D016022:Case-Control Studies; D002648:Child; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood; D007159:Immunologic Tests / Q000379:methods*; D007938:Leukemia / Q000469:parasitology* / Q000473:pathology; D015800:Protozoan Proteins / Q000032:analysis* / Q000097:blood / Q000235:genetics* / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014122:Toxoplasma / Q000235:genetics* / Q000276:immunology; D014123:Toxoplasmosis / Q000175:diagnosis* / Q000276:immunology / Q000473:pathology*
chemical_list: D000953:Antigens, Protozoan; C112573:GRA7 protein, Toxoplasma gondii; D007074:Immunoglobulin G; D015800:Protozoan Proteins; D011994:Recombinant Proteins
keywords: ELISA; LAMP; Patients with cancer; Recombinant dense granule protein 7; Toxoplasma gondii