Use of a recombinant Coccidioides immitis complement fixation antigen-chitinase in conventional serological assays

Abstract

The coccidioidal complement fixation (CF) antigen has been cloned previously, and the fusion protein has been expressed in Escherichia coli. The recombinant CF (rCF) antigen was affinity purified by adsorptiondesorption to chitin, and its reactivity was studied by using sera containing coccidioidal antibodies. The affinity-purified rCF antigen formed a line of identity with an immunodiffusion (ID) CF reference antigen (coccidioidin) derived from mycelial-phase Coccidioides immitis and was reactive with human, canine, and equine sera containing coccidioidal antibody. The affinity-purified rCF antigen yielded no detectable reaction with Blastomyces or Histoplasma antiserum by ID. The affinity-purified rCF antigen fixed complement with positive human sera and, even when used at lower concentrations, yielded titers comparable to those obtained with the coccidioidin. The reactivity of the affinity-purified rCF antigen was further evaluated by enzyme immunoassay, in which it manifested good sensitivity (96.9%) and specificity (100%) when evaluated with 43 human patients' sera. Thus, the affinity-purified rCF antigen has yielded reactions comparable to those of crude coccidioidal antigens in conventional CF, IDCF, and enzyme immunoassay.

Mapping of a Coccidioides immitis-specific epitope that reacts with complement-fixing antibody

“…Zimmerman et al (33), using oligonucleotide primers synthesized on the basis of the amino-terminal sequence of the native CF/chitinase protein, recently cloned the CF/chitinase protein, and in a subsequent study, Johnson et al (19) evaluated the utility of the recombinant protein in detecting anti-CF antibody when assayed by the CF test and an enzyme immunoassay. Their 47-kDa recombinant antigen was highly sensitive in detecting CF antibody by an enzyme immunoassay and the CF test.…”

Clinical Laboratory Utility of a Humanized Antibody in Commercially Available Enzyme Immunoassays for Coccidioidomycosis

“…The use of recombinant CTS1 (rCTS1) and truncations of rCTS1 have been tested to measure anti-CF antibodies in VF patient serum using an EIA format ( 15 , 17 , 18 ). Currently, there are two Food and Drug Administration (FDA)-approved EIAs used for the qualitative detection of anti-coccidioidal IgG and IgM, Premier Coccidioides EIA (Meridian Bioscience, Cincinnati, OH) and clarus Coccidioides antibody EIA (Immuno-Mycologics, Inc. [IMMY], Norman, OK).…”

Development of a quantitative antigen assay to detect coccidioidal chitinase-1 (CTS1) in human serum

“…CTS1 is also known as the "CF" antigen used in serodiagnostics, namely complement fixation (CF) assays and its immunodiffusion correspondent (IDCF) [16,17]. It has since been produced recombinantly by several groups, and truncations of CTS1 have been explored to map antibodybinding sites to increase sensitivity and specificity for detecting antibodies to Coccidioides in patients [19][20][21][22][23].…”