Production of in-vitro refolded and highly antigenic SAG1 for development of a sensitive and specific Toxoplasma IgG ELISA

Allahyari, Mojgan; Mohabati, Reyhaneh; Babaie, Jalal; Amiri, Samira; Siavashani, Zahra Jafari; Zare, Mehrak; Sadeghiani, Ghazaleh

doi:10.1016/j.jim.2014.11.012

Cited by 16 publications

(7 citation statements)

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“…Immunodominant surface antigen of T. gondii, SAG1, is debated a consequential antigen for progress of effective diagnostic tests and propose vaccines (19,20). SAG1 was selected in this study since in numerous studies it is reported as a good candidate for toxoplasma detection (13,18,(21)(22)(23)(24). Our results about the sensitivity and specificity of IgG-ELISA-rSAG1 was consistent with those reported by Khanaliha et al (22) and Jalallou et al (23).…”

Section: Discussionsupporting

confidence: 87%

Evaluation of Recombinant SRS3 Antigen for Diagnosis of Toxoplasmosis by Enzyme-Linked Immunosorbent Assay

Marashiyan

Moradian

Saadatnia

et al. 2016

Arch Clin Infect Dis

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Section: Discussionsupporting

confidence: 87%

Evaluation of Recombinant SRS3 Antigen for Diagnosis of Toxoplasmosis by Enzyme-Linked Immunosorbent Assay

Marashiyan

Moradian

Saadatnia

et al. 2016

Arch Clin Infect Dis

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“…HNP1 was synthesized with a purity of >90% (93.17%) by Biomatik (Cambridge, Canada) ( S1 Fig ) and then was folded by oxido-shuffling process [ 21 ]. Briefly, the purified HNP1 peptide at the concentration of 160 μg/ml was solubilized in folding buffer (Tris–HCL 50mM, NaCl 100mM, EDTA 0.1mM, reduced glutathione 3mM and oxidized glutathione 0.3 mM, pH 8.5) and kept at 4°C in rotator (40 rpm) for overnight, then diluted in 10X volume of changing buffer and centrifuged in 3000 × g at 4°C for 30 min to precipitate aggregates.…”

Section: Methodsmentioning

confidence: 99%

Human Neutrophil Peptide 1 as immunotherapeutic agent against Leishmania infected BALB/c mice

et al. 2017

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Human Neutrophil Peptide 1 (HNP1) produced by neutrophils, is a well-known antimicrobial peptide which plays a role both in innate as well as in adaptive immunity and is under intensive investigation as a potential therapeutic agent. Previous in vitro experiments have indicated the leishmaniacidal effect of recombinant HNP1 on Leishmania major (L. major) promastigotes and amastigotes. In the current study, we further extended the idea to explore the remedial effect of HNP1 in the two modalities of peptide therapy (folded HNP1) and gene therapy in L. major infected BALB/c mice. To this end, mice in five different groups received synthetic folded HNP1 (G1), pcDNA-HNP1-EGFP (G2), pcDNA-EGFP (G3), Amphotericin B (G4) and PBS (G5), which was started three weeks after infection for three consecutive weeks. Footpad swelling was monitored weekly and a day after the therapy ended, IFN-γ, IL-4, IL-10, IL-6 and nitric oxide produced by splenocytes were analyzed together with the parasite load in draining lymph nodes. Arginase activity and dermal histopathological changes were also analyzed in the infected footpads. We demonstrated that both therapeutic approaches effectively induced Th1 polarization and restricted parasite burden. It can control disease progression in contrast to non-treated groups. However, pcDNA-HNP1-EGFP is more promising in respect to parasite control than folded HNP1, but less effective than AmB treatment. We concluded with the call for a future approach, that is, a DNA-based expression of HNP1 combined with AmB as it can improve the leishmaniacidal efficacy.

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“…Recombinant SAG-1 was expressed in Escherichia coli cells as inclusion bodies (IBs). The IBs were separated from the cell extract by centrifugation, and washed using IB washing buffers, as previously described, 23 to remove bacterial impurities and cell debris. Subsequently, the IBs were solubilized using Tris-HCl buffer containing 8 M Urea, and rSAG-1 was purified by a single immobilized metal ion chromatography (IMAC – Bio-Rad Laboratories, Inc, USA).…”

Section: Methodsmentioning

confidence: 99%

Evaluation of Immunogenic Effect of Toxoplasma gondii RecombinantSAG-1 Antigen with Propranolol as Adjuvant in BALB/c Mice

et al. 2019

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Purpose: Propranolol as a novel adjuvant, was used to evaluate the immunogenic effect of three doses of recombinant SAG-1 (rSAG-1) antigen of Toxoplasma gondii in BALB/c mice for finding the optimal dose, and was compared with efficacy of tachyzoite lysate antigen (TLA). Methods: Eight different groups of 15 BALB/c mice received different volumes of the immunogenic material (three doses of r SAG-1 and one dose of TLA antigens), with or without propranolol adjuvant, subcutaneously. The control group mice received only PBS. Three weeks after the last immunization, the serum levels of IgG2a, IgG1 and IgG total antibodies against TLA, splenic interleukin-5 (IL-5) and Interferon-gamma (IFN-γ) (produced against TLA) and the splenic lymphocyte proliferation after adding TLA were measured to evaluate humoral and cellular immune responses. Challenge test was performed by subcutaneously injection of 1000 alive and active tachyzoites in to five mice per each group and survival days for each group of mice were recorded. Results: The mice group that received propranolol adjuvant and 20 µg of r SAG-1 antigen per dose of injection showed significantly more IFN-γ production, more proliferation of splenic lymphocytes and higher anti-TLA-specific IgG2a production (three main indexes for cell mediated immunity) in comparison with other groups. Moreover, in the challenge test, this group of mice had a significantly increased survival time, indicating the positive effect of propranolol in the more stimulating of cellular immunity that is necessary for toxoplasmosis prevention or suppress. Conclusion: Our results showed that T. gondii rSAG-1 antigen in combination with propranolol as adjuvant (which can induce Th1 related responses) are good candidates for further study to a vaccine design.

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Production of in-vitro refolded and highly antigenic SAG1 for development of a sensitive and specific Toxoplasma IgG ELISA

Cited by 16 publications

References 50 publications

Evaluation of Recombinant SRS3 Antigen for Diagnosis of Toxoplasmosis by Enzyme-Linked Immunosorbent Assay

Evaluation of Recombinant SRS3 Antigen for Diagnosis of Toxoplasmosis by Enzyme-Linked Immunosorbent Assay

Human Neutrophil Peptide 1 as immunotherapeutic agent against Leishmania infected BALB/c mice

Evaluation of Immunogenic Effect of Toxoplasma gondii RecombinantSAG-1 Antigen with Propranolol as Adjuvant in BALB/c Mice

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