However, IgG reactivities with MAG1, SAG1, and GRA8 showed different levels over times of infection

However, IgG reactivities with MAG1, SAG1, and GRA8 showed different levels over times of infection. These results were further confirmed by determination of the IgG antibody avidity for every single recombinant antigen. In the time course of infection, IgG antibodies against the early recognized antigens matured significantly earlier than those directed against the later antigens did. The IgA patterns did not give reliable information about the infection time points. The data revealed that the recombinant line assay provides valuable information on the actual state of infection, SB-334867 free base especially during the early infection SB-334867 free base time points. Toxoplasmosis is caused by the parasiteToxoplasma gondii. Cats are the primary host of this pathogen. Sexual reproduction takes place only in the primary host, leading to excretion of infectious oocytes that can be incorporated by humans while gardening or by the consumption of insufficiently washed fruits or vegetables. In an alternative pathway, humans can be infected as a result of consumption of insufficiently heated infected meat, sinceToxoplasma gondiipersists in pigs, goats, and other mammals. A further source of infection is diaplacental transmission ofToxoplasma gondiifrom an acutely infected mother to her unborn child (congenital toxoplasmosis). Congenital toxoplasmosis may cause abortion and serious damage to the fetus, with severe neurological disorders (reviewed in references27,29, and32). Therefore, an accurate diagnosis of toxoplasmosis during pregnancy and early treatment is crucial. Since the course of a toxoplasma infection is generally asymptomatic, the diagnosis is generally SB-334867 free base based on SB-334867 free base serological methods. The methods used are immunofluorescence, enzyme immunoassay, complement fixation, and immunosorbent agglutination assay (11,15,16). There are numerous test systems commercially available. In nearly all these test kits, various preparations of tachyzoite antigen that might be contaminated by nonparasitic material and that vary due to different antigen preparation methods are utilized. Thus, recombinantly producedToxoplasma gondiiantigens were considered to replace tachyzoite material in toxoplasmosis serology. In the past, a large number of different recombinant antigens were produced inEscherichia coliand studied for their potential to serve as diagnostic markers ofToxoplasma gondiiinfections; these included dense granule proteins GRA1 (p24 [1,2,5]), GRA2 (p28 [1,25]), GRA4 (p41 [1,20]), GRA6 (p32 [1,26]), GRA7 (p29 [1,2,10,12]), and GRA8 (p35 [1,2,12,13,31]); surface antigens SAG1 (p30 [1,2,4,7]) and SAG2 (p22 [1,22]); rhoptry antigens ROP1 (p66 [1,12]) and ROP2 (p54 [1,33]); matrix protein MAG1 (p65, p68 [1,12,23]); microneme proteins MIC3 and MIC5 (2); and other recombinant antigens ofToxoplasma gondii. In most studies the recombinant antigens were coated alone or in some cases in various combinations on enzyme-linked immunosorbent assay (ELISA) plates. A combination of GRA7, GRA8, and ROP1 was previously suggested for the detection ofToxoplasma gondii-specific immunoglobulin M (IgM) antibodies (1). A combination of GRA7, GRA8, and SAG1 (1) or of GRA7, GRA8, SAG2, and H4 (14) was proposed for the detection of IgG antibodies. The latter combination was supposed to be reactive in patients with acute profiles but not in patients with chronic profiles. So far, combinations of different recombinantly produced antigens have not been applied separately on nitrocellulose membranes. Such a technique would allow the detection of human anti-Toxoplasma gondiiantibodies directed against every single recombinant antigen as well as the determination of the HsT16930 avidities of the individual IgG antibodies. The main objective of all diagnostic efforts in toxoplasmosis serology (mostly as preventive measures during pregnancy) is clarification of whether or not the pregnant woman has been acutely infected or whether infection occurred before conception. Due to the fact that low IgM titers in most cases persist long beyond the acute phases of infection, confirmation of IgM antibodies in serum is an inadequate criterion for diagnosing an acute toxoplasmosis (17). Therefore, determination of the avidities of the IgG serum antibodies is a very important step in diagnostics (8,18). However, it has been shown that the IgG antibodies raised against the individualToxoplasmaantigen differ in their maturation characteristics (19,34). In particular, there were antigens found that did not induce the synthesis of high-avidity IgG antibodies at all (19,34). Thus, using the complete mixture of tachyzoite antigens as it is used in conventional avidity assays, the determination of avidity is compromised by these differences. The use of recombinant antigens in avidity determination might.