A novel igm‐indirect hemagglutination test for the serodiagnosis of acute toxoplasmosis

Abstract

A new indirect hemagglutination (M-Toxo-IHA) test was standardized for the serodiagnosis of acute toxoplasmosis. The reagent for this test consisted of stabilized human red cells coated with a Toxoplasma gondii heat-stable alkaline-solubilized extract which reacted predominantly with IgM antibodies found in serum samples from patients with a recent infection. Almost no reactivity was verified with serum samples from chronic or ancient toxoplasmosis. The reagent preparation was simple and obviated prior fractionation of parasite components reactive with IgM antibodies. Rheumatoid factor or high levels of IgG antibodies did not interfere in the M-Toxo-IHA test. A total of 358 serum samples was assessed for the evaluation of this test, comprising sera from patients with recent and ancient toxoplasmosis, from non-related diseases, as well as from clinically healthy individuals with no previous T. gondii infections. Serodiagnostic performance of the M-Toxo-IHA test in terms of sensitivity, specificity, efficiency, and positive and negative predictive values was verified, and the values obtained were 1.000, 0.985, 0.989, 0.952, and 1.000, respectively, in relation to IgM-immunofluorescence or IgM-capture ELISA. Six different batches of reagent, successively produced for the current study, provided reproducible results and were stable for at least 22 months at 4 degrees C.

Improvement of the indirect hemagglutination test for the detection of antibodies to Streptococcus pyogenes

“…The previous reagent was found to be stable only for 15 days at 4 o C, in contrast to the new reagent which lasted at least 9 months at 4 o C, 6 months at room temperature and 6 weeks at 37 o C. The processes to keep a reagent in liquid solutions without changing its reactivity are usually manufacturers patents or secret formulation. e) Serum dilutions with hypotonic saline solution (0.1 M NaCl) gave better results than the isotonic saline solution (0.15 M NaCl) used for the previous reagent, confirming the findings of Yamamoto et al (17) for the toxoplasma system; f) V-shaped plastic microplates showed clear-cut results within 90 min compared to U-shaped plates which gave results within 2 or 3 h. These conditions for the preparation of the ECP reagent were also optimal for obtaining the PSLO reagent.…”

“…The test was carried out with the standardized reagent in V-shaped wells of plastic microplates using 0.1 M NaCl for serum sample dilution, as described (17). Twenty-five microliters of sensitized cell suspension was added to 50 µl of serial double dilutions of serum samples from 1/20 to 1/5,120.…”

Detection of Toxoplasma gondii-Specific Antibodies in Dogs. A Comparative Study of Immunoenzymatic, Immunofluorescent and Haemagglutination Titers

“…Thus, a negative serological result generally rules out a diagnosis of toxoplasmosis and shows significance, as it may guide the procedures of the clinicians in research for other diseases; (2) seropositive samples in all three tests (20%) with high titers of antibodies and predominance of IgM antibodies detected mainly by the M-Toxo test. This test, developed for the serodiagnosis of acute human toxoplasmosis (Yamamoto et al 1991), was carried out with the dog serum samples in order to verify its suitability for detection of IgM-specific antibodies in dogs. It was found 4 positive samples by M-Toxo out of 8 positive samples (50%) for IgG antibodies in all three tests, suggesting that these animals might be presenting an active infection by T. gondii.…”

“…All positive serum samples in the IHA test were retested after treatment with 2-Mercaptoethanol (2-ME) in order to verify the presence of IgM antibodies (Camargo et al 1978). Other IHA (M-Toxo) developed by Yamamoto et al (1991) for the serodiagnosis of acute toxoplasmosis was carried out with a standardized suspension of red blood cells coated with a heat-stable alkaline-solubilized extract of T. gondii, which reacts predominantly with IgM antibodies. To 50 µl of doubling diluted serum, as above mentioned, 25 µl of sensitized cell suspensions were added and the agglutination pattern read after incubation of 1 hr and 30 min at room temperature.…”

The Laboratory Diagnosis in Toxoplasma Infection

“…The result is considered the lecture at the higher dilution. Afterwards Yamamoto in 1991, standardized a test for the determination of IgM antibodies, with a sensibility of 98% and speciicity of 95% [16].…”