An immunocapture assay was developed to detect Toxoplasma gondii-specific immunoglobulin E (IgE) in sera from adults with acute acquired infection or reactivation and from babies with congenital toxoplasmosis. The components of this assay were monoclonal antibody to human IgE, samples from patients, and T. gondii tachyzoites treated with Formalin. When T. gondu-specific IgE antibodies were present, visually detectable agglutination occurred. Sera, umbilical cord blood, fetal blood, cerebrospinal fluid, and amniotic fluid were tested by this method. Specific IgE antibodies were detected in sera from 25 (86%) of 29 adults who developed specific IgG antibody during pregnancy or had specific IgA and IgM antibodies. Specific IgE was present early during infection, at the time that IgM antibodies were present, and slightly preceding the presence of specific IgA antibodies. In 23 patients tested serially, IgE antibodies never persisted for longer than 4 months. No nonspecific anti-T. gondii IgE was detected in sera from uninfected individuals. Maternal IgE antibodies did not cross the placenta. In sera of patients with congenital toxoplasmosis, specific IgE antibodies were found at birth, during the first year of life, and during immunologic recrudescence following discontinuation of pyrimethamine-sulfonamide therapy. The IgE immunocapture assay is simple to perform. It is especially useful for determining when T. gondu was acquired by recently infected pregnant women.

Enzyme-linked immunofiltration assay (ELIFA) for labeling transferred proteins is an interesting and powerful technique for the rapid specific detection (15 min) of proteins immobilized on nitrocellulose or nylon membranes (0.20 and 0.45 micron). ELIFA does not require fastidious handling of the membranes. Saturation, specific labeling and washing procedures are achieved by filtration, controlled by a monitoring unit which regulates the flow rate and ensures excellent specificity, repetition and reproducibility. The recycling by closed circuit or by repetitive inversion of the flow direction offers the advantage of reducing the volumes of expensive reagents while simultaneously increasing the sensitivity of the technique. The detection limit is at least as low as 1-5 ng using directly or indirectly enzymatically labelled probes. ELIFA may be extended to the identification of glycoproteins using specific ligands such as lectins or to the immunocapture of an antigen using specific antibodies immobilized on an activated membrane. ELIFA complements fast separation, by e.g., isoelectric focusing, polyacrylamide gel electrophoresis, or sodium dodecyl sulfate-polyacrylamide gel electrophoresis and accelerated electrotransfer to membranes with rapid detection reducing the total time for separation transfer and detection to less than 2 h.

To determine their prognostic and diagnostic values for toxoplasmosis in immunodepressed subjects, we assayed immunoglobulin A (IgA) and IgE antibodies by means of immunocapture (IC) tests, with revelation done by using a suspension of T. gondii (ICT). We also carried out a simultaneous analytical study of IgG antibodies on cellulose acetate membranes by using the comparative immunological profile method and an enzyme-linked immunofiltration assay (ELIFA). A total of 1,238 samples (serum, cerebrospinal fluid, and aqueous humor from 318 patients) were tested. IgA and IgE antibodies were detected in all heart, kidney, and liver transplant recipients with clinical manifestations of toxoplasmosis; IgA was detected in the aqueous humor of a patient with chorioretinitis. In patients with AIDS-related toxoplasmosis, including the cerebral form, IgA and IgE antibodies or a significant modification of ELIFA IgG values were observed in 38, 19, and 25% of patients, respectively. IgM was detected by ICT only in 12% of patients and aided the diagnosis in 1 of 71 patients. IC tests for specific IgA and IgE alone and combined with ELIFA were positive in 39 and 46% of patients who developed clinical toxoplasmosis, respectively. In a serial study of 16 patients in whom at least one of these three tests was positive, a significant immunological signal sometimes preceded clinical onset by 1, 6, and even 17 months. Similarly, in a group of human immunodeficiency virus-infected patients with evidence of previous exposure to T. gondii but no clinical manifestations, IgA, IgE, and IgA and/or IgE antibodies were detected in only 11, 4, and 12% of patients, respectively. These two situations point to peripheral T. gondii reactivation. IgA and IgE emerged as interesting markers of the risk of toxoplasmosis in immunodepressed patients. They may also provide valuable assistance in the diagnosis of toxoplasmosis, especially because tests for specific IgM are disappointing. However, at least one in two patients with toxoplasmosis showed no detectable immunological reaction, suggesting that this polyisotypic approach should be combined with other noninvasive methods such as gene amplification.

The target antigens of specific immunoglobulins G, M, A, and E from patients with acquired acute toxoplasmosis were determined using immunocytochemistry. The relative repartition of these antigens in four cellular compartments of Toxoplasma (membrane complex, apical area, rhoptries, and dense granules) was quantitatively evaluated. Rhoptry antigens mainly react positively with IgA. Membrane, submembrane area (membrane complex), and rhoptry antigens are immunodominant for IgA and IgM. Apical area antigens are recognized by IgM two times more than IgG and IgA. IgE recognized only rhoptry antigens. The localization of pathogenetically antigenic components and their identification by the immune system appeared to be of importance for selection of immunodominant or recombinant antigens. Such localization would improve laboratory diagnosis of serious congenital toxoplasmosis or in immunocompromised patients with toxoplasmic complications after cyst reactivation.