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2.
Am J Clin Pathol ; 85(6): 704-9, 1986 Jun.
Article in English | MEDLINE | ID: mdl-3706207

ABSTRACT

Optimum methodologic variables for assessing cellular immunity by in vitro lymphocyte transformation (LT) were determined using spherulin and coccidioidin antigens. This study was conducted in an area endemic for coccidioidomycosis and included healthy, coccidioidomycosis skin test positive (STP) and negative (STN) subjects, and patients with mild, acute disease. The authors examined the relationship between coccidioidin (1:100) and spherulin (low dose) skin test reactivity and lymphocyte transformation (LT) responses to the same antigens. Counts per minute (CPM) and stimulation index (SI) as methods of expressing tritiated thymidine uptake were compared. The LT assays were set up in duplicate test systems using autologous and homologous plasma. Both antigens differentiated between STP and STN groups (P less than or equal to 0.001-0.004), but values obtained with spherulin-induced LT were greater than those using coccidioidin (P less than 0.001). Values in CPM and SI were greater in the spherulin-induced LT assay using autologous compared with AB plasma. Specifically, for detecting cellular immunity to coccidioidomycosis, the combination of spherulin-induced LT using autologous plasma and expressing the results in CPM gave the best discrimination between STP and STN subjects. Based on epidemiologic data, the latter method also appeared more sensitive than the skin test in detecting cellular immunity to coccidioidomycosis. In general, these data illustrate the variable effectiveness of different antigens for inducing LT responses and further show how different plasma sources affect the LT response. Finally, these data suggest that lymphocyte blast transformation results expressed as CPM may give more consistent values and better discrimination between immune and nonimmune subjects than results expressed as stimulation indices.


Subject(s)
Lymphocyte Activation , Adult , Coccidioidin/analysis , Coccidioidomycosis/immunology , Evaluation Studies as Topic , Female , Humans , Immunity, Cellular , Male , Methods
3.
Am J Trop Med Hyg ; 33(4): 645-9, 1984 Jul.
Article in English | MEDLINE | ID: mdl-6476210

ABSTRACT

Clinical findings in patients with primary infection due to Coccidioides immitis have suggested that fungal antigenemia may occur. To study this possibility, we devised an inhibition enzyme-linked immunosorbent assay (ELISA) to measure antigenic activity compared to the activity of antigens derived from spherules. Twenty-one of 27 sera from 19 infected patients showed antigenic activity above that found in sera from uninfected persons. Activity was highest early in the course of illness. In other studies, anti-spherulin antibody activity was measured by ELISA, which demonstrated an association of antigen activity with IgM, but not IgG or IgA. One possible explanation for this phenomenon is that IgM antibody and antigen are present as circulating immune complexes.


Subject(s)
Antibodies, Fungal/analysis , Antigens, Fungal/analysis , Coccidioidin/analysis , Coccidioidomycosis/immunology , Adolescent , Adult , Coccidioidin/immunology , Enzyme-Linked Immunosorbent Assay , Humans , Immunoglobulin A/analysis , Immunoglobulin G/analysis , Immunoglobulin M/analysis , Male , Middle Aged , Time Factors
4.
Infect Immun ; 23(2): 479-85, 1979 Feb.
Article in English | MEDLINE | ID: mdl-422249

ABSTRACT

The interpretation of immunological results in systemic mycoses has been complicated by cross-reactions among specimens from patients with blastomycosis, coccidioidomycosis, and histoplasmosis. The fungal preparations used in these tests evidently contained one or more antigens in common. Two-dimensional immunoelectrophoresis techniques were used to determine the number of antigens contained in several soluble extracts from Blastomyces dermatitidis and Histoplasma capsulatum that were common with those demonstrable in a coccidiodin-anticoccidioidin reference system. A total of 12 and 10 common antigens were found in preparations from B. dermatitidis and H. capsulatum, respectively. In addition, the crude preparations from each species of fungus exhibited some qualitative and quantitative differences in composition of antigens. Use of two-dimensional immunoelectrophoresis for standardization of fungus extracts, for monitoring separation of species-specific antigens, and for testing humoral antibody response should add further refinement to correlations with clinical disease.


Subject(s)
Antigens, Fungal/analysis , Blastomyces/immunology , Coccidioides/immunology , Histoplasma/immunology , Coccidioidin/analysis , Immunoelectrophoresis
5.
Infect Immun ; 20(2): 541-51, 1978 May.
Article in English | MEDLINE | ID: mdl-669811

ABSTRACT

Immunological tests are valuable aids for diagnosis of mycotic infections and, in some cases, as objective guides for clinical management and prognosis. The usefulness of these procedures is limited to the extents that crude antigen preparations are employed, that these are difficult to standardize uniformly, and that they contain antigens common to several species of pathogenic fungi. Analysis by two-dimensional immunoelectrophoresis methods of the two crude preparations used for coccidioidomycosis demonstrated that coccidioidin contained at least 26 antigens, with 10 of these found also in spherulin. In addition, spherulin contained two antigens not demonstrated in coccidioidin. No single test detected all antigens present, and multiple procedures were required to display the complete array of antigens. A reference system was established for coccidioidin and precipitated immunoglobulins from a burro hyperimmunized with coccidioidin. Evaluation of the reference system demonstrated that it was highly reproducible with respect to the reagents used, to repeated tests by the same person, and to comparative tests by two individuals using the same reagents. Applications of this reference system for standardization of reagents, for detecting common antigens, for monitoring successive steps during fractionation of crude preparations, and for fingerprinting strains for ecological and epidemiological studies are presented.


Subject(s)
Antigens, Fungal/analysis , Coccidioides/immunology , Coccidioidin/analysis , Coccidioidomycosis/immunology , Immunoelectrophoresis, Two-Dimensional/methods
6.
J Clin Microbiol ; 5(2): 149-53, 1977 Feb.
Article in English | MEDLINE | ID: mdl-66238

ABSTRACT

An immunological procedure for the rapid and specific identification of Coccidioides immitis isolates has been developed. The specificity of the procedure is based on the fact that C. immitis produces antigens that are not produced by morphologically similar fungi. The procedure involved the transfer of heavy mold-form inocula to flasks that contained small volumes of brain heart infusion broth. Shake cultures were grown at 25 degrees C on a gyratory shaker at 150 rpm. The concentrated supernatant antigens so obtained from broth cultures were tested in parallel with coccidioidin by a microimmunodiffusion technique against C. immitis antiserum. The ability of the immunological procedure to identify C. immitis cultures was evaluated by testing the supernatant antigens of 166 unknown isolates, many of which, by gross or microscopic examination or both, resembled C. immitis or belonged to the family Gymnoascaceae. Each culture was also identified by conventional laboratory procedures. Comparative evaluation showed that the immunological test for C. immitis was 100% sensitive and specific. The diagnostic precipitin bands that were produced by the interaction of the C. immitis supernatants and antisera were shown to consist of (i) a heat-stable antigen comparable to that active in the tube precipitin test and (ii) two heat-labile antigens, one of which was associated with complement fixation reactivity. With pure cultures, this immunological procedure permitted the identification of C. immitis isolates within 5 days.


Subject(s)
Antigens, Fungal/analysis , Coccidioides/isolation & purification , Immunodiffusion , Coccidioides/immunology , Coccidioidin/analysis , Epitopes , Evaluation Studies as Topic , Humans
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