Toxoplasma gondiiinfection: analysis of serological response by 2-DE immunoblotting

Abstract

Toxoplasma gondii is known to cause a variety of diseases ranging from asymptomatic infections to serious conditions in immunocompromised hosts such as AIDS-patients or transplant recipients. In addition they may cause abortion or fetal abnormalities during pregnancy. Despite the clinical importance, diagnosis, treatment and prevention still remain unsatisfactory. Analysis of the parasitic cell determinants, recognized by specific humoral and cellular immune responses, may have important implications for diagnosis, therapy and vaccination strategies. Two-dimensional electrophoresis (2-DE) was used to resolve and compare protein patterns from Toxoplasma gondii strains RH and BK (mouse virulent strains). Comparison of silver-stained gels showed that 35.2% to 60.3% of the spots had the same position. In a second series of experiments, the reactivity of the spots with human sera was tested. Proteins were transferred to PVDF membranes and were detected with sera from different patient groups. Depending upon the immunoglobulin class (IgG, IgM, IgA or IgE) different epitope patterns were observed. Some of the spots seemed to be recognized in different stages of infection. Sera of two patients with similar serology and comparable stage of infection were compared in order to demonstrate an individual immune response.

Application of proteomics for comparison of proteome of Neospora caninum and Toxoplasma gondii tachyzoites

“…Moreover, 2-DE combined with immunoblotting assay is allowed to find out many and distinct antigens compared with conventional SDS-PAGE (one-dimensional) and its immunoblotting analysis [29,30]. These methods have been employed successfully in characterizing the antigen profiles of T. gondii either with monoclonal antibody or patient's sera [13,14]. 2-DE map of T. gondii was reported [12] and detected average 630 (591-685) spots from 2-DE gel using pH 4-7 IPG strips (18 cm) stained with silver nitrate.…”

“…The two-dimensional gel electrophoresis with powerful image analysis software and biological mass spectrometry in combination with database searching made it possible to analyze complex protein mixtures extracted from cells, tissues, or other biological samples [10,11]. These proteomic methods have been proved successfully for characterizing the proteome of T. gondii [12] and 2-DE combined with immunoblotting assay enables characterizing the antigen profiles of T. gondii using specific antibodies [13,14]. The comparison of 2-DE antigen profiles between N. caninum and T. gondii has been conducted by using specific antisera [15], however, they could not identify the antigen spots that showing different 2-DE profiles between them.…”