Preparation of Aspergillus Fumigatus Antigens and Their Analysis by Two-Dimensional Immunoelectrophoresis

Hearn, Veronica M.; Wilson, Elaine V.; Proctor, Andrew; Mackenzie, D. W. R.

doi:10.1099/00222615-13-3-451

Cited by 47 publications

(40 citation statements)

References 4 publications

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“…'s method of two-dimensional immunoelectrophoresis is used. This is corroborated by more recent studies of Hearn et al [16].…”

Section: Aspergillus Mycosissupporting

confidence: 81%

Significance of Immunological Methods for Diagnosis of Fungal Disease

Seeliger¹

1982

Chemotherapy

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In the diagnosis of classical systemic mycoses, as caused by Blastomyces dermatidis, Coccidioides immitis, Histoplasma capsulatum and Paracoccidioidesbrasiliensis, serodiagnostic procedures in conjunction with skin tests are of significant diagnostic and prognostic value. The diagnosis of aspergillus mycosis is facilitated greatly by serological tests and the same is true for recognition of cryptoeoccus mycosis by means of serological detection of the cryptococcal antigen. Great progress has also been made in the evaluation of Candida antibodies in patients’ sera.

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“…'s method of two-dimensional immunoelectrophoresis is used. This is corroborated by more recent studies of Hearn et al [16].…”

Section: Aspergillus Mycosissupporting

confidence: 81%

Significance of Immunological Methods for Diagnosis of Fungal Disease

Seeliger¹

1982

Chemotherapy

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“…However, the results obtained by these methods have been inconsistent due to differences in the sensitivity of the detection methods and the lack of standardization of antigen preparations. Indeed, antigen extracts have been made from A. fumigatus cultures incubating for as short as 3 [12,13,21] to as long as 2 weeks after incubation [23]. Furthermore, material has been prepared from culture filtrates [12], and from mechanical [II, 13,21] or chemical extrac tions [23] from fungal cultures.…”

Section: Discussionmentioning

confidence: 99%

“…Indeed, antigen extracts have been made from A. fumigatus cultures incubating for as short as 3 [12,13,21] to as long as 2 weeks after incubation [23]. Furthermore, material has been prepared from culture filtrates [12], and from mechanical [II, 13,21] or chemical extrac tions [23] from fungal cultures. The absence of stan dardization has caused large variations in the antigen icity of commercial and noncommercial lots.…”

Section: Discussionmentioning

confidence: 99%

Localization, Molecular Weight and Immunoglobulin Subclass Response to <i>Aspergillus fumigatus</i> Allergens in Acute Bronchopulmonary Aspergillosis

Leung

Gershwin

Coppel

et al. 1988

Int Arch Allergy Immunol

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A specimen of Aspergillus fumigatus was isolated from a patient with acute bronchopulmonary aspergillosis (ABPA). Cultures were allowed to propagate and were separated into spores and mycelium to > 95% homogeneity. Extracts of both the spores and mycelium were prepared and used for study by both dot blot analysis and immunoblotting to study the major allergens. By dot blot analysis, 22 of 22 patients with ABPA and none of 10 healthy controls reacted with mycelium when probed for both IgG and IgE anti-Aspergillus reactivity. Similar results were obtained when these extracts were separated on polyacrylamide gel electrophoresis and then probed; this included both IgG and IgE reactivity. Further, the reactivity to preparations of whole extract was absorbed with mycelium but not spores. Several distinct protein bands were detected with IgG, ranging from 30 to 110 kilodaltons (kD). In contrast, the majority of patients with ABPA, when studied for IgE reactivity, reacted only with a 70-kD protein, although 1 out of 22 patients reacted with a 30-kD protein. Subclass analysis of IgG reactivity demonstrated that 90% of reactive sera contained IgG2 and 73% IgG4 reactivity. IgG1 and IgG3 reactivity were present but less frequently detected. The use of immunoblotting will enable the identification of relevant allergens. Further, the demonstration that the allergenicity is located primarily in mycelium will allow more definitive studies of allergen isolation for studies of reactivity and, ultimately, cloning of the 70-kD protein and, finally, epitope mapping.

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“…Tests were done on 8.2 x 7.6 cm slides in a 1.2 mm layer of agarose gel 1% (w/v), buffered with Tris barbitone at pH 8.6 (Hearn et al, 1980). The antigen well held 7-25 p1 of solution at the cathodic end of the plate.…”

Section: Methodsmentioning

confidence: 99%

Use of Aspergillus fumigatus mycelial antigens in enzyme-linked immunosorbent assay and counter-immunoelectrophoresis

Wilson

Hearn

1983

Journal of Medical Microbiology

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SUMMARY. Antigens obtained from the ruptured mycelium of Aspergillus fumigatus were separated on the basis of their attachment or non-attachment to Concanavalin A-linked Sepharose gel. They were analysed by double-diffusion and two-dimensional immunoelectrophoresis with rabbit antiserum and sera from patients with aspergillus-related disease. Their sensitivity was assessed in an enzyme-linked immunosorbent assay (ELISA) for immunoglobulin G. Sera which gave very low or high extinction readings on ELISA gave, in general, comparable results on counter-immunoelectrophoresis; but weak electrophoretic reactions were not closely paralleled by ELISA results. The unfractionated extract and the fraction bound by Concanavalin A-Sepharose had similar ability to detect specific IgG in sera from patients with aspergillosis.

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Preparation of Aspergillus Fumigatus Antigens and Their Analysis by Two-Dimensional Immunoelectrophoresis

Cited by 47 publications

References 4 publications

Significance of Immunological Methods for Diagnosis of Fungal Disease

Significance of Immunological Methods for Diagnosis of Fungal Disease

Localization, Molecular Weight and Immunoglobulin Subclass Response to <i>Aspergillus fumigatus</i> Allergens in Acute Bronchopulmonary Aspergillosis

Use of Aspergillus fumigatus mycelial antigens in enzyme-linked immunosorbent assay and counter-immunoelectrophoresis

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