Isolation of Coccidioides immitis F antigen by immunoaffinity chromatography with monospecific antiserum

Cox, Rebecca A.; Britt, L A; Michael, R A

doi:10.1128/iai.55.1.227-232.1987

Cited by 20 publications

(10 citation statements)

References 16 publications

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“…Immunoblots of CDN and spherulin (8) probed with polyclonal goat anti-CF, the murine anti-CF MAb, and CF-positive sera from coccidioidomycosis patients yielded a 43-to 45-kDa couplet band (8). Although the size of the CF couplet band detected in our studies was lower than that reported by Zimmer and Pappagianis (14), both the 43-to 45-kDa and 48-to 50-kDa couplet bands were shown to be labile to heat treatment (56ЊC, 30 min) and proteolytic enzymes but resistant to periodate oxidation or treatment with glycolytic enzymes (6,8,14), indicating that they are the same proteins. This conclusion is further supported by the fact that both we (8) and Zimmer and Pappagianis (14) showed that the couplet bands represented a product of a native 110-kDa protein, as evidenced by the detection of the latter as opposed to the couplet bands when blotting was performed with gels that had been electrophoresed under nondenaturing, nonreducing conditions.…”

Section: Discussioncontrasting

confidence: 67%

“…Towards this goal, Zimmer and Pappagianis (31) identified the CF antigen as a 48-kDa peptide (subsequently characterized as a 48-to 50-kDa couplet [14]) by immunoblotting with CF antibody-positive coccidioidomycosis sera. Concomitant with those studies, we identified the CF antigen by comparing the CF activity and antigenic composition of CDNderived fractions in two-dimensional immunoelectrophoresis against a hyperimmune goat anti-CDN and, by using the precipitin arc as an immunogen, developed a monospecific goat anti-CF (6). In a subsequent study, Dolan and Cox (8) developed a murine IgG1 MAb which recognizes a peptide epitope that is specific to the CF antigen.…”

Section: Discussionmentioning

confidence: 99%

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Molecular cloning and characterization of the Coccidioides immitis complement fixation/chitinase antigen

et al. 1996

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Detection of anti-Coccidioides complement-fixing (CF) antibody is a valuable diagnostic and prognostic aid in coccidioidomycosis. The CF antibody response is directed against a heat-labile antigen that has chitinase activity, hereafter referred to as the CF/chitinase protein. To identify and clone this immunoreactive enzyme, we constructed a Coccidioides immitis cDNA lambda ZAP expression library from spherule RNA and detected fusion peptides expressing CF epitopes by immunoscreening. A cDNA clone consisting of 1,623 bp was identified, sequenced, and found to contain a single open reading frame that encodes a protein of 47 kDa with 427 amino acids. Deduced amino acid sequence analyses showed that the cloned CF/chitinase cDNA contains a 35-amino-acid region, beginning at Ser-18 and ending at Arg-52, which has 92% homology with the reported N-terminal amino acid sequence of authentic CF/chitinase protein. The first 17 amino acids in the deduced sequence of the cloned cDNA are not present on the mature CF/chitinase protein, suggesting that it may be a signal peptide. Expression of the CF/chitinase cDNA insert by using the pGEX-4T-3 vector yields a fusion peptide that bears CF-specific epitopes and shows chitinase activity. The CF/chitinase clone will enable large-scale production of the recombinant CF antigen for use in immunoassays and facilitate studies on the role of chitinase in the morphogenesis of C. immitis.

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confidence: 67%

Section: Discussionmentioning

confidence: 99%

Molecular cloning and characterization of the Coccidioides immitis complement fixation/chitinase antigen

et al. 1996

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“…The important diagnostic and prognostic value of the CF antibody is well established (16, 28, 29). Naturally, the corresponding CF antigen has generated considerable interest and research (6,13,24,34). When it was demonstrated that antibody detected by IDCF correlated with antibody detected by CF, the antigen producing the precipitate in the IDCF test was assumed to be the same antigen which participated in CF (13).…”

Section: Discussionmentioning

confidence: 99%

The coccidioidal complement fixation and immunodiffusion-complement fixation antigen is a chitinase

Johnson

Pappagianis

1992

Infect Immun

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Culture filtrates and autolysates of Coccidioides immitis have provided suitable crude antigens for the serodiagnosis and prognosis of coccidioidomycosis. One of these, a heat-labile antigen which participates in the immunodiffusion reaction corresponding to the complement fixation reaction (IDCF), has been characterized as a 110-kDa native protein that, when subjected to reducing conditions and heat, yields a 48-kDa component. The present report provides serologic and biochemical evidence that this antigen is a chitinase. This chitinase, isolated from 48-h culture filtrate of the spherule-endospore-phase C. immitis by affinity adsorption to chitin, formed a line of identity with the IDCF reference antigen and participated in the complement fixation reaction with human serum. It lost its enzymatic as well as antigenic activity when heated, but when not heated it retained its enzymatic activity even when precipitated with coccidioidal antibody present in human serum. This chitinase represents a significant serodiagnostic substance and may be important in the morphogenesis of C. immitis.

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“…This component was further characterized by using size exclusion chromatography followed by ID and denaturing SDS-PAGE, as well as nondenaturing PAGE, and shown to be derived from a 110-kDa protein nonreactive with lentil lectin or concanavalin A (158). Cox et al (30) isolated an antigen reactive with IDCF antiserum by using a combination of concanavalin A affinity and immunoaffinity chromatography. This antigen was assayed by immunoelectrophoresis and corresponded to antigen 3 in the reference system of Huppert et al (66).…”

Section: Antigen Characterizationmentioning

confidence: 99%

Serology of coccidioidomycosis.

Pappagianis¹,

Zimmer²

1990

Clinical Microbiology Reviews

100

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and, more recently, enzyme-linked immunosorbent assay (ELISA). NATURE OF C. IMMITIS C. immitis exists in nature and in the usual culture media in its saprobic form: septate hyphae 2 to 4 Rm in diameter which, in 5 to 7 days, yield a chain of multinucleate arthroconidia which usually alternate with smaller, nonviable, brittle cells (Fig. 1). The latter degenerate, thus releasing the arthroconidia, which readily become airborne. The arthroconidia can germinate to yield new hyphae or can serve as the form infecting humans and other hosts. When inhaled, the arthroconidia, in the presence of the phagocytic cells (42) and increased CO2 (80), convert into a different morphologic form. Shedding an outer wall layer and all but one nucleus, the arthroconidia round up and enlarge to produce an immature spherule. The nucleus undergoes division, which is followed by partitioning of the cytoplasm by inward extension of the cell wall. Completion of the nuclear division and segmentation produce a mature spherule with endospores within 48 h in vivo and at 30 to 96 h in vitro. Following the segmentation, the endospores become

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Isolation of Coccidioides immitis F antigen by immunoaffinity chromatography with monospecific antiserum

Abstract: Detection of antibody to Coccidioides immitis F antigen is of proved value in the diagnosis of coccidioidomycosis. This antibody is demonstrable by use of an immunodiffusion assay with reference

Cited by 20 publications

References 16 publications

Molecular cloning and characterization of the Coccidioides immitis complement fixation/chitinase antigen

Molecular cloning and characterization of the Coccidioides immitis complement fixation/chitinase antigen

The coccidioidal complement fixation and immunodiffusion-complement fixation antigen is a chitinase

Serology of coccidioidomycosis.

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