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    Toxoplasma Serology Laboratory:
    A Guide for Clinicians

    Laboratory Tests for the Diagnosis of Toxoplasmosis

    Table of Contents

    The diagnosis of toxoplasmosis may be established by serological tests, polymerase chain reaction (PCR), histological demonstration of the parasite and/or its antigens (i.e. immunoperoxidase stain), or isolation of the organism. The Toxoplasma Serology Laboratory of the Palo Alto Medical Foundation Research Institute (TSL-PAMFRI) offers confirmatory serological and PCR testing and methods for isolation of the organism. In addition, our medical consultants (Jose G. Montoya, M.D., Jack S. Remington, M.D. and Despina Contopoulos-Ioannidis, M.D.) are available for interpretation of test results and advice on management of patients.

    Serological Tests

    Different serological tests often measure different antibodies that possess unique patterns of rise and fall with time after infection. A combination of serological tests is frequently required to establish whether an individual has been more likely infected in the distant past or has been recently infected.

    Brain Tissue Sample
    Brain tissue at autopsy of an AIDS patient with toxoplasmic encephalitis.
    Wright Giemsa stain of cerebrospinal fluid demonstrating T. gondii tachyzoites.
    T. gondii cyst
    Scanning microscopy of a T. gondii cyst.
    Acute toxoplasmosis in eyeFundoscopic examination of an AIDS patient with ocular toxoplasmosis.
    An IgM test is used to help determine whether a patient has been infected recently or in the distant past. Because of the significant potential of misinterpreting a positive IgM test result, confirmatory testing should be performed. Despite the wide distribution of commercial test kits to measure IgM antibodies, these kits often have low specificity and the reported results are frequently misinterpreted. In addition, IgM antibodies can persist for months to more than one year.

    Toxoplasma Serological Profile (TSP)

    For confirmatory testing, TSL-PAMFRI offers a panel of tests (the Toxoplasma Serological Profile (TSP) comprised of the Sabin-Feldman Dye Test (DT), double sandwich IgM ELISA, IgA ELISA, IgE ELISA, and AC/HS test. The TSP has been used successfully by our group to determine whether serological test results are more likely consistent with infection acquired in the recent or more distant past.

    The TSP has been clinically helpful in the setting of toxoplasmic lymphadenitis, myocarditis, polymyositis, chorioretinitis, and during pregnancy. For sera with positive results in IgG and IgM tests, the discriminatory power of the TSP to differentiate between recently acquired and chronic infection is superior to any single serological test.

    Recently, several tests for avidity of toxoplasma IgG antibodies have been introduced to help discriminate between recently acquired and distant infection. Studies of the kinetics of the avidity of IgG in pregnant women who have seroconverted during gestation have shown that women with high avidity test results were infected with T. gondii at least 3 to 5 months earlier (time to conversion from low to high avidity antibodies varies with the method used). Because low avidity antibodies may persist for many months, their presence does not necessarily indicate recently acquired infection.

    IgG Antibodies

    Dye Test

    At TSL-PAMFRI IgG antibodies are primarily measured by the Sabin-Feldman Dye Test (DT). The DT is a sensitive and specific neutralization test in which live organisms are lysed in the presence of complement and the patient's IgG T. gondii -specific antibody. IgG antibodies usually appear within 1 to 2 weeks of the infection, peak within 1 to 2 months, fall at variable rates, and usually persist for life. The titer does not correlate with the severity of illness.

    A positive DT establishes that the patient has been exposed to the parasite. A negative DT essentially rules out prior exposure to T. gondii (unless the patient is hypogammaglobulinemic). However, in a small number of patients, IgG antibodies might not be detected within 2 to 3 weeks after the initial exposure to the parasite. In addition, rare cases of toxoplasmic chorioretinitis and TE (toxoplasmic encephalitis) in immunocompromised patients have been documented in patients negative for T. gondii -specific IgG antibodies.

    Differential agglutination (AC/HS)

    The differential agglutination test (also known as the "AC/HS test") uses two antigen preparations that express antigenic determinants found early following acute infection (AC antigen) or in the later stages of infection (HS). Ratios of titers using AC versus HS antigens are interpreted as acute, equivocal, non-acute patterns of reactivity or non-reactive. The acute pattern may persist for one or more years following infection. This test has proved useful in helping differentiate acute from chronic infections but is best used in combination with a panel of other tests (e.g.: the TSP).


    The functional affinity of specific IgG antibodies is initially low after primary antigenic challenge and increases during subsequent weeks and months. Protein-denaturing reagents including urea are used to dissociate the antibody-antigen complex. The avidity result is determined using the ratios of antibody titration curves of urea-treated and untreated serum.

    We routinely employ the avidity test as an additional confirmatory diagnostic tool in the TSP for those patients with a positive and/or equivocal IgM test or acute and/or equivocal pattern in the AC/HS test. Health care providers and clinical laboratories involved in the care of pregnant women should be aware that avidity testing is a confirmatory test and not the ultimate test for decision-making. Its highest value is observed when laboratory test results reveal high IgG avidity antibodies and the serum is obtained during the time window of exclusion of acute infection for a particular method (i.e. 12 weeks for the Labsystems method, 16 weeks for the VIDAS-bioMérieux method). Low or equivocal IgG avidity antibody results should not be interpreted as diagnostic of recently acquired infection. These low or equivocal avidity antibodies can persist for months to one year or longer.

    IgM Antibodies

    IgM antibodies are measured by the "double-sandwich" or "immuno-capture" IgM-ELISA method. This method avoids false positive results due to the presence of rheumatoid factor and antinuclear antibodies.

    In patients with recently acquired infection, IgM T. gondii antibodies are detected initially and, in most cases, these titers become negative within a few months. However, in some patients, positive IgM T.gondii -specific titers can be observed during the chronic stage of the infection. IgM antibodies have been reported to persist as long as 12 years after the acute infection. Persistence of these IgM antibodies does not appear to have any clinical relevance and these patients should be considered chronically infected.

    The FDA has recommended that sera with positive IgM test results obtained at non-reference laboratories should be sent to a Toxoplasma reference laboratory. At our reference laboratory, these referred IgM positive sera undergo confirmatory testing and the results are interpreted as i) a recently acquired infection, ii) an infection acquired in the distant past or iii) a false positive result.

    IgA Antibodies

    IgA antibodies may be detected in sera of acutely infected adults and congenitally infected infants using ELISA or ISAGA methods. As is true for IgM antibodies to the parasite, IgA antibodies may persist for many months to more than one year. For this reason they are of little additional assistance for diagnosis of the acute infection in the adult. In contrast, the increased sensitivity of IgA assays over IgM assays for diagnosis of congenital toxoplasmosis represents an advance in diagnosis of the infection in the fetus and newborn. In a number of newborns with congenital toxoplasmosis and negative IgM antibodies, the serological diagnosis has been established by the presence of IgA and IgG antibodies.

    IgE Antibodies

    IgE antibodies are detectable by ELISA in sera of acutely infected adults, congenitally infected infants, and children with congenital toxoplasmic chorioretinitis. The duration of IgE seropositivity is less than with IgM or IgA antibodies and hence appears useful as an adjunctive method for identifying recently acquired infections.

    Polymerase Chain Reaction (PCR)

    * See the What's New page for additional information.

    PCR amplification is used to detect T. gondii DNA in body fluids and tissues. It has been successfully used to diagnose congenital, ocular, cerebral and disseminated toxoplasmosis. PCR performed on amniotic fluid has revolutionized the diagnosis of fetal T. gondii infection by enabling an early diagnosis to be made, thereby avoiding the use of more invasive procedures on the fetus. PCR has allowed detection of T. gondii DNA in brain tissue, cerebrospinal fluid (CSF), vitreous and aqueous fluid, bronchoalveolar lavage (BAL) fluid, urine, amniotic fluid and peripheral blood.

    Histologic Diagnosis (Testing Not Performed At Our Facility)

    Demonstration of tachyzoites in tissue sections or smears of body fluid (e.g., CSF, amniotic fluid or BAL) establishes the diagnosis of the acute infection. It is often difficult to demonstrate tachyzoites in conventionally stained tissue sections. The immunoperoxidase technique, which uses antisera to T. gondii , has proven both sensitive and specific; it has been successfully used to demonstrate the presence of the parasite in the central nervous system of AIDS patients. The immunoperoxidase method is applicable to unfixed or formalin-fixed paraffin-embedded tissue sections. We do not perform the immunoperoxidase method but we assist in the interpretation of already stained slides or facilitate performance of the stain at the Department of Pathology at Stanford University School of Medicine.

    A rapid and technically simple method is the detection of T. gondii in air-dried, Wright-Giemsa-stained slides of centrifuged (e.g., cytocentrifuge) sediment of CSF or of brain aspirate or in impression smears of biopsy tissue. Our laboratory can assist in reviewing such preparations.

    The presence of multiple tissue cysts near an inflammatory necrotic lesion probably establishes the diagnosis of acute infection or reactivation of latent infection.

    Isolation of T. gondii

    Isolation of T. gondii from blood or body fluids establishes that the infection is acute. Attempts at isolation of the parasite can be performed at TSL-PAMFRI by mouse inoculation.