1Associate Professor, Department of Pathobiology, School of Veterinary Medicine, Shahid Bahonar University of Kerman, Kerman, Iran
2Department of Pathobiology, School of Veterinary Medicine, Shahid Bahonar University of Kerman, Kerman, Iran
3Associate Professor, Department of Parasitology, School of Veterinary Medicine, Shahid Bahonar University of Kerman, Kerman, Iran
Background & Aims: Toxoplasma gondii is an obligate, intracellular parasite, which is widely spread in the world. The parasite is able to infect all warm-blooded hosts including humans and farm animals. The infection in humans often occurs after the ingestion of raw or undercooked meat containing tissue cysts. Several methods have been applied to detect this parasite in contaminated foods. Recombinant antibodies are new generation of monoclonal antibodies, which are isolated via phage display technology from immune or non-immune phage libraries against target antigens. These antibodies are used for diagnosis of many different antigens and therapeutics proposes. The object of the present study was to isolate recombinant monoclonal antibodies against this important parasite. Methods: The purified Toxoplasma gondii surface antigen, P30, was coated to immunotubes and used as a target for selection of antibodies from the Tomlinson I and J phage display libraries of single-chain fragment variable (scFv) antibodies. Clones that were able to recognize antigen were isolated in three rounds of binding, elution and amplification. The specificity of scFv antibodies chosen from the resulting panel, were confirmed using enzyme-linked immunosorbent assay (ELISA), dot blotting and western blotting methods. Results: Recombinant antibodies capable of recognizing P30 antigen were isolated with high affinity; and their specificity was approved. Conclusion: Isolated soluble single chain antibodies are good candidates to apply as monoclonal recombinant antibodies in diagnostic kits for detection of Toxoplasma gondii in contaminated samples.
Montoya JG. Laboratory diagnosis of Toxoplasma gondii infection and toxoplasmosis. J Infect Dis 2002; 185(Suppl 1): S73-S82.
Ogouyemi-Hounto A, Agbayahoun-Chokki F, Sissinto Savi de TY, Biokou BB, Adinsi dS, V, Assogba M, et al. [Evaluation of a rapid diagnostic test in the diagnosis of toxoplasmosis in pregnant women in Cotonou (Benin)]. Bull Soc Pathol Exot 2014; 107(2): 85-9.
Chong CK, Jeong W, Kim HY, An DJ, Jeoung HY, Ryu JE, et al. Development and clinical evaluation of a rapid serodiagnostic test for toxoplasmosis of cats using recombinant SAG1 antigen. Korean J Parasitol 2011; 49(3): 207-12.
Hoogenboom HR, de Bruinea AP, Hufton SE, Hoet RM, Arends JW, Roovers RC. Antibody phage display technology and its applications. Immunotechnology 1998; 4(1): 1-20.
Bradbury AR, Marks JD. Antibodies from phage antibody libraries. J Immunol Methods 2004; 290(1-2): 29-49.
Sommavilla R, Lovato V, Villa A, Sgier D, Neri D. Design and construction of a naive mouse antibody phage display library. J Immunol Methods 2010; 353(1-2): 31-43.
Tomlinson I&J libraries protocol. Available at: Http://www.geneservice.co.uk/products/proteomic/datasheets/tomlinsonIJ.pdf.
Caravelli A, Luz DE, Andrade FB, Moraes CT, Maranhao AQ, Piazza RM. Sensitive and specific detection of enteropathogenic and enterohemorrhagic Escherichia coli using recombinant anti-intimin antibody by immunofluorescence assay. Diagn Microbiol Infect Dis 2013; 77(4): 301-3.
Hairul Bahara NH, Tye GJ, Choong YS, Ong EB, Ismail A, Lim TS. Phage display antibodies for diagnostic applications. Biologicals 2013; 41(4): 209-16.
Weisser NE, Hall JC. Applications of single-chain variable fragment antibodies in therapeutics and diagnostics. Biotechnol Adv 2009; 27(4): 502-20.
Sambrook J, Russell DW. Molecular cloning: a laboratory manual. New York, NY: CSHL Press; 2001.
Munoz-Zanzi CA, Fry P, Lesina B, Hill D. Toxoplasma gondii oocyst-specific antibodies and source of infection. Emerg Infect Dis 2010; 16(10): 1591-3.
Cook AJ, Gilbert RE, Buffolano W, Zufferey J, Petersen E, Jenum PA, et al. Sources of toxoplasma infection in pregnant women: European multicentre case-control study. European Research Network on Congenital Toxoplasmosis. BMJ 2000; 321(7254): 142-7.
Zia-Ali N, Fazaeli A, Khoramizadeh M, Ajzenberg D, Darde M, Keshavarz-Valian H. Isolation and molecular characterization of Toxoplasma gondii strains from different hosts in Iran. Parasitol Res 2007; 101(1): 111-5.
Asgari Q, Sarnevesht J, Kalantari M, Sadat SJ, Motazedian MH, Sarkari B. Molecular survey of Toxoplasma infection in sheep and goat from Fars province, Southern Iran. Trop Anim Health Prod 2011; 43(2): 389-92.
Ergin S, Ciftcioglu G, MIdilli K, Issa G, Gargili A. Detection of toxoplasma gondii from meat and meat products by the nested-pcr method and its relationship with seroprevalence in slaughtered animals. Bull Vet Inst Pulawy 2009; 53: 657-61.
Aspinall TV, Marlee D, Hyde JE, Sims PF. Prevalence of Toxoplasma gondii in commercial meat products as monitored by polymerase chain reaction--food for thought? Int J Parasitol 2002; 32(9): 1193-9.
Dubey JP, Hill DE, Jones JL, Hightower AW, Kirkland E, Roberts JM, et al. Prevalence of viable Toxoplasma gondii in beef, chicken, and pork from retail meat stores in the United States: risk assessment to consumers. J Parasitol 2005; 91(5): 1082-93.
Golchin M, Khalili-Yazdi A, Karamouzian M, Abareghi A. Latex agglutination test based on single-chain Fv recombinant antibody fragment. Scand J Immunol 2012; 75(1): 38-45.
Spadiut O, Capone S, Krainer F, Glieder A, Herwig C. Microbials for the production of monoclonal antibodies and antibody fragments. Trends Biotechnol 2014; 32(1): 54-60.