Reevaluation of commercial reagents for detection of Histoplasma capsulatum antigen in urine.

Detection of the Histoplasma capsulatum urinary antigen (UAg) is among the most sensitive and rapid means to diagnose histoplasmosis. Previously, we evaluated analyte-specific reagents (ASR) manufactured by IMMY (Norman, OK) for detection of Histoplasma galactomannan (GM) in urine using an enzyme immunoassay (EIA), and we showed low positive agreement (64.5%) with the MiraVista (MVista) Histoplasma antigen (Ag) quantitative EIA (MiraVista Diagnostics, Indianapolis, IN). Here we reevaluated the IMMY GM ASR following modification of our original assay protocol and introduction of an indeterminate range. A total of 150 prospectively collected urine samples were tested with both the IMMY and MVista EIAs, and clinical histories were recorded for all study subjects. The IMMY GM ASR showed positive and negative agreements of 82.3% (14/17 samples) and 100% (121/121 samples), respectively (with exclusion of 12 indeterminate results), and overall agreement of 90% (135/150 samples) with respect to the MVista EIA. Of the three patients with negative IMMY GM ASR results and positive MVista EIA results, testing was performed for initial diagnostic purposes for one patient (<0.4 ng/ml by the MVista EIA) and UAg levels were being monitored for the remaining two patients (both<0.7 ng/ml by the MVista EIA). The MVista EIA results were positive for 6/12 samples that tested indeterminate by the IMMY GM ASR. We also show that the IMMY GM ASR can be used to serially monitor Histoplasma UAg levels. In conclusion, we demonstrate that, with modification, the IMMY GM ASR is a reliable rapid assay for detection of Histoplasma UAg.

Detection of the dengue virus NS1 antigen using an enzyme immunoassay.

Current diagnostic methods for dengue virus (DV) rely primarily on detection of anti-DV antibodies and/or DV RNA by reverse transcriptase (RT) PCR. Several limitations exist however: seroconversion is delayed following infection, and DV RT-PCR assays are not yet readily available. The DV nonstructural protein 1 (NS1) antigen is an alternative acute phase DV biomarker, and here, we evaluated the new InBios (InBios International, Inc., Seattle, WA, USA) DENV Detect(TM) NS1 enzyme-linked immunoassay (ELISA) compared to DV RT-PCR and serology for detection of recent DV infection. We report a positive, negative, and overall percent agreement of 96% (24/25), 86.0% (43/50), and 89.3% (67/75) for the InBios NS1 ELISA compared to DV RT-PCR. Performance of the NS1 ELISA compared to serology for anti-DV IgM antibodies showed a positive, negative, and overall percent agreement of 78.0% (85/109), 90.7% (333/367), and 87.8% (418/476), respectively. Collectively, the InBios NS1 ELISA can be used as an alternative to DV RT-PCR for identification of acute DV infection.

Detection of (1, 3)-ß-D-glucan in bronchoalveolar lavage and serum samples collected from immunocompromised hosts.

The incidence of invasive fungal infections (IFI) has increased in recent years, especially among immunocompromised hosts (ICH). In 2003, the Fungitell(®) assay received FDA clearance for the presumptive diagnosis of IFI using serum and detects (1-3)-ß-D-glucan, which is a major cell wall component of certain fungi (e.g., Candida, Aspergillus, and Pneumocystis). The goal of the current study was to assess the performance of the assay on bronchoalveolar lavage (BAL) fluid and serum to identify IFI in ICH. Patients were classified as having proven, probable, possible, or no IFI according to the European Organization for Research and Treatment of Cancer/Mycoses Study Group (EORTC/MSG) guidelines. Among 109 patients for whom the results of Fungitell were compared to the EORTC/MSG criteria, Fungitell showed a low positive predictive value for the identification of IFI from both BAL (20.0%) and serum (26.7%). However, the negative predictive value of Fungitell was significantly higher for both sample types (BAL, 83.0%; serum, 84.8%). Interestingly, the results of Fungitell were positive in BAL and serum in 7/8 (87.5%) patients diagnosed with Pneumocystis pneumonia (PcP) by real-time, non-nested PCR. These data indicate that the Fungitell assay has a low positive predictive value for the diagnosis of IFI in ICH, regardless of the specimen type that is tested. However, testing of serum samples by Fungitell may permit a rapid and noninvasive initial screening approach in patients with presumed PcP.

Direct comparison of the traditional and reverse syphilis screening algorithms in a population with a low prevalence of syphilis.

We describe the first direct comparison of the reverse and traditional syphilis screening algorithms in a population with a low prevalence of syphilis. Among 1,000 patients tested, the results for 6 patients were falsely reactive by reverse screening, compared to none by traditional testing. However, reverse screening identified 2 patients with possible latent syphilis that were not detected by rapid plasma reagin (RPR).

Evaluation of the Bio-Rad BioPlex Measles, Mumps, Rubella, and Varicella-Zoster Virus IgG multiplex bead immunoassay.

The goal of this study was to compare the BioPlex 2200 measles, mumps, rubella, and varicella-zoster virus (MMRV) IgG multiplex assays (Bio-Rad Laboratories, Hercules, CA) to routine testing by enzyme immunoassay (EIA). Serum specimens (n = 500) submitted to our reference laboratory for routine MMRV IgG testing by EIA were also tested by the BioPlex assays. Following testing, the BioPlex measles, mumps, rubella, and varicella-zoster virus assays demonstrated agreements of 91.6% (95% confidence interval [CI], 88.8% to 93.7%), 94.2% (95% CI, 91.7% to 95.7%), 94.4% (95% CI, 92.0% to 96.1%), and 91.8% (95% CI, 89.0% to 93.9%), respectively, compared to the results of EIA. Timing studies showed that the BioPlex MMRV assay could provide complete analysis of 100 serum specimens in 1.7 h, compared to 5.5 h by EIA. These data indicate that the BioPlex MMRV IgG assays exhibit comparable performance (93% overall agreement [1,860/2,000 results]; κ = 0.67) to routine testing by EIA. The BioPlex assays allow for the simultaneous detection of all four analytes, thereby eliminating potential aliquot errors and reducing turnaround time.

Detection of IgG-class antibodies to measles, mumps, rubella, and varicella-zoster virus using a multiplex bead immunoassay.

Serologic testing for measles, mumps, rubella, and varicella (MMRV) IgG is traditionally performed by immunofluorescence assay or enzyme immunoassay (EIA). Although sensitive and specific, these methods are labor intensive, time consuming, and require separate assays for each analyte. This study evaluated the performance of the MMRV IgG AtheNA Multi-Lyte assay using nonclinically characterized serum specimens submitted to our laboratory for routine MMRV IgG testing. Mumps (n = 492) or rubella (n = 500) IgG were initially tested by enzyme-linked fluorescent antibody (ELFA), whereas measles (n = 494) or varicella (n = 497) were analyzed by EIA. Each sample was also tested by the AtheNA Multi-Lyte assay. Discordant results were retested by the predicate method and the multiplex assay, with further discrepancies being arbitrated by a third test. Compared to EIA/ELFA for MMRV IgG, the AtheNA assay demonstrated an overall agreement of 97.4%, 98.2%, 97.6%, and 100%, respectively. Use of this multiplex assay allows for the simultaneous detection of MMRV IgG, potentially decreasing cost, sample volume requirements, aliquot errors, and hands-on testing time.