The second-generation Histoplasma antigen immunoassay is semiquantitative, expressing results as a comparison to a negative control, which requires repeat testing of the prior specimen with the current specimen to accurately determine a change in antigen. Reporting results in this manner often is confusing to the ordering physician and laboratory. Development of a quantitative assay could improve accuracy, reduce interassay variability, and eliminate the need to test the prior sample with the current sample in the same assay. Calibrators with known concentrations of Histoplasma antigen were used to quantitate antigen in specimens from patients with histoplasmosis and from controls. Samples from cases of disseminated histoplasmosis or other mycoses and controls were tested to evaluate the performance characteristics of the quantitative assay. Paired specimens were evaluated to determine if quantitation eliminated the need to test the current and prior specimens in the same assay to assess a change in antigen. The sensitivity in samples from patients with AIDS and disseminated histoplasmosis was 100% in urine and 92.3% in serum. Cross-reactions occurred in 70% of other endemic mycoses, but not in aspergillosis. Specificity was 99% in controls with community-acquired pneumonia, medical conditions in which histoplasmosis was excluded, or healthy subjects. A change in antigen level categorized as an increase, no change, or decrease based on antigen units determined in the same assay agreed closely with the category of change in nanograms/milliliter determined from testing current and prior specimens in different assays. Sensitivity, specificity, and interassay precision are excellent in the new third-generation quantitative Histoplasma antigen immunoassay.Antigen detection is a useful method for the diagnosis of histoplasmosis (8). Until now, antigen results have been expressed semiquantitatively as antigen units (calculated as enzyme immunoassay [EIA] units [EU]) based upon comparison to a negative control. Antigen levels decline with effective therapy (9, 12) and increase with relapse (13), providing a useful method for monitoring treatment. Based upon analysis of the course of antigen clearance during treatment of histoplasmosis in patients with AIDS (4, 6, 7), a Ն4-EU increase in antigen was chosen as evidence suggesting a relapse of histoplasmosis. In that evaluation, the change in antigen units between current and prior specimens tested concurrently rarely exceeded 4 EU in patients who did not relapse clinically (unpublished observation).Due to interassay variability, prior specimens have been tested simultaneously with current specimens to accurately assess changes in antigen levels. Test reports including current and prior results often are confusing, however, and do not always reach the ordering physician. In this report, a method for quantitation of Histoplasma capsulatum antigen is described, and its performance characteristics, including accuracy, specificity, sensitivity in patients with AIDS and dissemi...

We observed false-positive results in the Platelia Aspergillus enzyme-linked immunoassay (EIA) for specimens from patients with histoplasmosis and mice with experimental infection. Platelia Aspergillus EIA-positive specimens were negative in the second-generation Histoplasma antigen EIA. Care must be taken to exclude histoplasmosis for patients with positive Platelia Aspergillus EIA results.The Platelia Aspergillus enzyme-linked immunoassay (EIA) detects a galactomannan antigen produced by several molds (5). However, studies to date have not included Histoplasma capsulatum. We recently observed false-positive results in the Platelia Aspergillus EIA for specimens from six patients with culture-proven histoplasmosis, as have others (4). Based upon these observations (Table 1), we conducted a laboratory-based study of specimens submitted for Platelia Aspergillus EIA or Histoplasma antigen testing and evaluated cross-reactivity in experimental models of histoplasmosis and aspergillosis.(Part of this work was presented at the 46th Interscience Conference on Antimicrobial Agents and Chemotherapy, 27 to 30 September 2006.)Residual serum and bronchoalveolar lavage (BAL) fluid specimens that were submitted to MiraVista Diagnostics for Histoplasma antigen testing or Platelia Aspergillus EIA and were positive were retested the following day in the other EIA. The second-generation Histoplasma antigen EIA (MiraVista Diagnostics, Indianapolis, IN) uses polyclonal antibodies to H. capsulatum and has been described elsewhere (6). Results of Ն1 unit were regarded as positive. The Platelia Aspergillus EIA (Bio-Rad Laboratories, Redmond, WA) uses monoclonal antibodies produced against Aspergillus fumigatus. Specimens were pretreated with EDTA for the Platelia Aspergillus EIA and boiled in accordance with the manufacturer's specifications. Results with a galactomannan index (GMI) of 0.5 or greater were reported as positive. Note that the Platelia Aspergillus EIA is not FDA cleared for specimens other than serum.Twenty-three of 48 serum specimens positive for antigen in the second-generation Histoplasma antigen EIA were positive in the Platelia Aspergillus EIA (Fig. 1). Positive results were more frequent for specimens giving levels of 40 units or higher in the Histoplasma antigen EIA (12/17 [70.6%]) than for those giving levels below 40 units (11/31 [35.5%]) (P ϭ 0.043 by chi-square test). As controls, 12 serum specimens that were negative in the Histoplasma antigen EIA were tested in the Platelia Aspergillus EIA, and all were negative. Seven of 11 (63.6%) BAL fluid specimens that were positive in the Histoplasma antigen EIA were positive in the Platelia Aspergillus EIA. Results for the Histoplasma antigen EIA ranged from 2.2 to 61.7 units for the BAL fluid specimens that were positive in the Platelia Aspergillus EIA, compared to 4.2 to 21.2 units for those that were negative. Ten control BAL fluid specimens that were negative in the Histoplasma antigen EIA were negative in the Platelia Aspergillus EIA.Twenty serum specimens that w...