Development of a Quantitative Antigen Assay to Detect Coccidioidal Chitinase-1 (CTS1)

Abstract

Background Coccidioidomycosis is often diagnosed with a collection of tests that rely on the patient’s ability to mount an immune response to the fungus (antibody-based diagnostics), making diagnosis of this infection challenging. Here we present an antigen-based assay that detects and quantifies coccidioidal chitinase-1 (CTS1) in human serum. Methods An inhibition-based enzyme-linked immunoassay (ELISA) was developed that utilizes a monoclonal antibody specific for coccidioidal CTS1. CTS1 was quantified in commercial antigen preparations using recombinant CTS1 as a standard. Sera from 192 individuals from an endemic area were tested which included 78 patients (40.6%) with proven or probable coccidioidomycosis. Results The quantity of CTS1 in diagnostic commercial antigen preparations from different suppliers varied. CTS1 antigenemia was detected in 87.2% of patients with proven or probable coccidioidomycosis. Specificity was determined to be 96.94% using serum from individuals who reside in the Phoenix, Arizona area who did not have coccidioidomycosis. Levels of CTS1 correlated with low- and high-titer serology from patients with a coccidioidomycosis diagnosis. Conclusions Since the CTS1 inhibition ELISA described in this report does not depend on the host immune response, it is a promising diagnostic tool to aid in diagnosis and disease monitoring of coccidioidomycosis.

Persistent Headaches in an Avid Hiker: A Case of Chronic Coccidioidal Meningitis

“…A case series of three patients with refractory CM showed clinical improvement with isavuconazole, used as salvage therapy [ 15 ]. Prompt serologic CM diagnosis through Coccidioides antigen testing, serum and CSF antibodies by immunodiffusion and complement fixation, and a promising new coccidioidal chitinase-1 (CTSI) inhibition enzyme-linked immunosorbent assay (ELISA) can aid in timely treatment to prevent clinical deterioration [ 16 ].…”

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

“…The amount of CTS1 in commercial antigen preparations used for immunodiffusion was quantified using our previously published assay ( 20 ) in order to normalize the amount of CTS1 between antigens. Based on the amount quantified in each antigen, 40 ng of CTS1 was prepared with 4× sodium dodecyl-sulfate (SDS) sample loading buffer for subsequent SDS-polyacrylamide gel electrophoresis (PAGE).…”

“…This requirement has therefore highlighted the need for consistent IgG and IgM dilution controls for clinical laboratories performing Coccidioides EIAs. While the composition and concentration of the antigens used in both commercial EIAs are proprietary information, previous data generated by our laboratory have demonstrated that CTS1 is a component present in complement fixation and immunodiffusion antigens supplied by IMMY and Meridian ( 20 ). This finding further suggests that CTS1 is a component of the antigenic preparation used in each EIA, whether intentional or not.…”

Immunologic Diagnosis of Endemic Mycoses

“…Specificity was determined to be 97% using sera from Phoenix, Arizona residents who did not have coccidioidomycosis. Levels of CTS1 could be associated with low- and high-titer serology from individuals with proven coccidioidomycosis diagnosis [ 50 ]. The CTS1 antigen detection assay has the possibility of similar or better performance than other immunologic assays as well as the distinct advantage of a direct measurement of fungal antigen concentrations in blood.…”