Deborah J. Jespersen

Treponema-Specific Tests for Serodiagnosis of Syphilis: Comparative Evaluation of Seven Assays

ABSTRACT The diagnosis of syphilis is challenging and often relies on serologic tests to detect treponemal or nontreponemal antibodies. Recently, the Centers for Disease Control and Prevention and the Association of Public Health Laboratories proposed an update to the syphilis serology testing algorithm, in which serum samples are first tested using a treponema-specific test and positive samples are analyzed with a nontreponemal assay. The goal of this study was to compare the performance of seven treponemal assays (BioPlex 2200 syphilis IgG [Bio-Rad, Hercules, CA], fluorescent treponemal antibody [FTA] assay [Zeus Scientific, Raritan, NJ], Treponema pallidum particle agglutination [TP-PA; Fujirebio Diagnostics, Malvern, PA], Trep-Sure enzyme immunoassay [EIA; Phoenix Biotech, Oakville, Ontario, Canada], Trep-Chek EIA [Phoenix Biotech], Trep-ID EIA [Phoenix Biotech], and Treponema ViraBlot IgG [Viramed Biotech AG, Planegg, Germany]) using serum samples (n = 303) submitted to our reference laboratory. In addition to testing with these 7 assays, all samples were tested by a rapid plasma reagin (RPR) assay and a treponemal IgM Western blot assay (Viramed ViraBlot). Compared to the FTA assay as the gold standard, the evaluated treponemal tests demonstrated comparable levels of performance, with percent agreement ranging from 95.4% (95% confidence interval, 92.3 to 97.3) for the Trep-Sure EIA to 98.4% (96.1 to 99.4) for the Trep-ID EIA. Compared to a “consensus of the test panel” (defined as at least 4 of 7 treponemal tests being in agreement), the percent agreement ranged from 95.7% (92.7 to 97.5) for Trep-Sure to 99.3% (97.5 to 99.9) for Trep-ID. These data may assist clinical laboratories that are considering implementing a treponemal test for screening or confirmatory purposes.

Direct Comparison of the Traditional and Reverse Syphilis Screening Algorithms in a Population with a Low Prevalence of Syphilis

ABSTRACT 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).

Comparison of four assays for the detection of cryptococcal antigen.

ABSTRACT We compared the performance of four assays for the detection of cryptococcal antigen in serum samples (n = 634) and cerebrospinal fluid (CSF) samples (n = 51). Compared to latex agglutination, the sensitivity and specificity of the Premier enzyme immunoassay (EIA), Alpha CrAg EIA, and CrAg lateral flow assay (LFA) were 55.6 and 100%, 100 and 99.7%, and 100 and 99.8%, respectively, from serum samples. There was 100% agreement among the four tests for CSF samples, with 18 samples testing positive by each of the assays.

Evaluation of a Multiplex Flow Immunoassay for Detection of Epstein-Barr Virus-Specific Antibodies

ABSTRACT Conventional methods for the detection of Epstein-Barr virus (EBV)-specific antibodies include the immunofluorescence assay (IFA) and enzyme immunoassay (EIA). While sensitive and specific, these methods are labor-intensive and require separate assays for each analyte. This study evaluated the performance of a multiplex bead assay (BioPlex 2200; Bio-Rad Laboratories, Hercules, CA) for the simultaneous detection of immunoglobulin G (IgG) and IgM class antibodies to the EBV viral capsid antigen (VCA) and IgG class antibodies to Epstein-Barr virus nuclear antigen-1 (EBNA-1). Serum specimens (n = 1,315) submitted for routine EBV-specific antibody testing by EIA (Grifols-Quest, Inc., Miami, FL) were also tested by the multiplex bead assay using the BioPlex 2200 automated analyzer. Specimens showing discordant results were tested by IFA. Following IFA resolution, the BioPlex VCA IgM, VCA IgG, and EBNA-1 IgG assays demonstrated 97.9%, 91.4%, and 96.9% agreement, respectively, with the results obtained by EIA. Furthermore, the BioPlex assays showed an overall agreement of 94.1% with the EIA when the specimens were categorized by disease state (susceptible, acute, or past infection) based on the EBV-specific antibody profiles. These findings indicate that the BioPlex EBV assays demonstrate a performance comparable to that of the conventional EIA, while allowing for a more rapid (2.3 h for 100 samples versus 4.5 h by the EIA) and higher-throughput (∼400 samples per 9 h versus 200 samples by the EIA) analysis of the EBV-specific antibody response.

Evaluation of the Bio-Rad BioPlex 2200 Syphilis Multiplex Flow Immunoassay for the Detection of IgM- and IgG-Class Antitreponemal Antibodies

ABSTRACT The laboratory diagnosis of syphilis is based primarily upon serologic findings. Historically, serologic testing for syphilis has relied on assays such as rapid plasma reagin, fluorescent treponemal antibody absorption, Treponema pallidum particle agglutination (TP-PA), and more recently, enzyme immunoassay (EIA). In this study, we evaluated the performance of a novel multiplex flow immunoassay (BioPlex 2200 Syphilis; Bio-Rad Laboratories, Hercules, CA) for the detection of antitreponemal IgG- and IgM-class antibodies. Serum specimens (n = 1,008) submitted for routine treponema-specific antibody testing by syphilis IgM and IgG EIA (Trep-Chek; Phoenix-Biotech, Mississauga, Ontario, Canada) were also analyzed by the BioPlex Syphilis multiplex assay. Specimens showing discordant results were repeat tested, with further discrepancies being arbitrated by TP-PA. Compared directly to the results of EIA, the BioPlex IgG assay demonstrated 98.7% (77/78) sensitivity and 99.4% (916/930) specificity. Compared to the Trep-Chek IgM EIA, the BioPlex IgM assay showed 80% (4/5) sensitivity and 97.9% (652/666) specificity. These results indicate that the BioPlex Syphilis multiplex assay shows similar serological agreement with EIA while allowing for a fully automated random-access platform that provides faster (1.7 h for 100 samples versus 4.5 h by EIA) and higher-throughput (800 samples per 9 h versus 200 samples by EIA) analysis of the syphilis serologic response.

Evaluation of three commercial latex agglutination kits and a commercial enzyme immunoassay for the detection of cryptococcal antigen.

We compared the performance of the Meridian CALAS, Wampole Crypto-LA, Murex Cryptococcus latex agglutination assay, and the Meridian Premier EIA for the detection of cryptococcal antigen in serum and CSF. The assays demonstrated similar performance characteristics based on concordance values > or = 93% but important differences were noted in endpoint titers.

Evaluation of Two Commercial Systems for Automated Processing, Reading, and Interpretation of Lyme Borreliosis Western Blots

ABSTRACT The diagnosis of Lyme borreliosis (LB) is commonly made by serologic testing with Western blot (WB) analysis serving as an important supplemental assay. Although specific, the interpretation of WBs for diagnosis of LB (i.e., Lyme WBs) is subjective, with considerable variability in results. In addition, the processing, reading, and interpretation of Lyme WBs are laborious and time-consuming procedures. With the need for rapid processing and more objective interpretation of Lyme WBs, we evaluated the performances of two automated interpretive systems, TrinBlot/BLOTrix (Trinity Biotech, Carlsbad, CA) and BeeBlot/ViraScan (Viramed Biotech AG, Munich, Germany), using 518 serum specimens submitted to our laboratory for Lyme WB analysis. The results of routine testing with visual interpretation were compared to those obtained by BLOTrix analysis of MarBlot immunoglobulin M (IgM) and IgG and by ViraScan analysis of ViraBlot and ViraStripe IgM and IgG assays. BLOTrix analysis demonstrated an agreement of 84.7% for IgM and 87.3% for IgG compared to visual reading and interpretation. ViraScan analysis of the ViraBlot assays demonstrated agreements of 85.7% for IgM and 94.2% for IgG, while ViraScan analysis of the ViraStripe IgM and IgG assays showed agreements of 87.1 and 93.1%, respectively. Testing by the automated systems yielded an average time savings of 64 min/run compared to processing, reading, and interpretation by our current procedure. Our findings demonstrated that automated processing and interpretive systems yield results comparable to those of visual interpretation, while reducing the subjectivity and time required for Lyme WB analysis.

Multiplex Detection of IgM and IgG Class Antibodies to Toxoplasma gondii, Rubella Virus, and Cytomegalovirus Using a Novel Multiplex Flow Immunoassay

ABSTRACT The goal of this study was to evaluate the BioPlex 2200 Toxoplasma, rubella, and cytomegalovirus (CMV) (ToRC) IgG and IgM multiplex immunoassays (Bio-Rad Laboratories, Hercules, CA) and compare the results to those of conventional testing by enzyme immunoassay (EIA) and enzyme-linked fluorescent assay (ELFA). Serum specimens (n = 600) submitted for routine ToRC IgG and IgM testing by EIA (SeraQuest, Doral, FL; Diamedix, Miami, FL) or ELFA (Vidas; bioMérieux, Durham, NC) were also tested by the BioPlex ToRC multiplex immunoassays. Samples showing discordant results were retested by both methods, with further discrepancies being arbitrated by a third assay. Following repeat testing, the BioPlex Toxoplasma, rubella, and CMV IgG assays demonstrated agreements of 98.7 (592/600 specimens), 93.3 (560/600 specimens), and 98.3% (590/600 specimens), respectively, while the ToRC IgM assays yielded agreements of 91.2 (547/600 specimens), 87.3 (524/600 specimens), and 95.2% (571/600 specimens), respectively. The BioPlex ToRC IgG assays provided results comparable to EIA/ELFA results, with kappa coefficients showing near-perfect agreement for the Toxoplasma (κ = 0.94) and CMV (κ = 0.97) IgG assays and substantial agreement for the rubella IgG assay (κ = 0.66). The BioPlex ToRC IgM assays showed lower specificity with only slight agreement for Toxoplasma IgM (κ = 0.07), poor agreement for rubella IgM (κ = −0.03), and moderate agreement for CMV IgM (κ = 0.55). Both the BioPlex IgG and IgM assays reduced turnaround time (1.7 h versus 5.5 h by EIA/ELFA for 100 specimens) and eliminated the necessity to manually pipette or aliquot specimens prior to testing.

Prospective Comparison of Cell Cultures and Nucleic Acid Amplification Tests for Laboratory Diagnosis of Chlamydia trachomatis Infections

ABSTRACT Specimens submitted in M5 medium for cell culture detection of Chlamydia trachomatis were tested by nucleic acid amplification testing (NAAT) and in cell cultures. Of 35 (genital) and 26 (nongenital) specimens positive for C. trachomatis, 21 and 14 specimens, respectively, were detected exclusively by NAAT. NAAT is significantly (P < 0.0001) more sensitive than cell culture and should be considered the new “gold standard” for the laboratory diagnosis of C. trachomatis infections.

Comparison of Three Immunoassays for Detection of Antibodies to Strongyloides stercoralis

ABSTRACT Due to the limited sensitivities of stool-based microscopy and/or culture techniques for Strongyloides stercoralis, the detection of antibodies to this intestinal nematode is relied upon as a surrogate for determining exposure status or making a diagnosis of S. stercoralis infection. Here, we evaluated three immunoassays, including the recently released InBios Strongy Detect IgG enzyme-linked immunosorbent assay (ELISA) (InBios International, Inc., Seattle, WA), the SciMedx Strongyloides serology microwell ELISA (SciMedx Corporation, Denville, NJ), and the luciferase immunoprecipitation system (LIPS) assay performed at the National Institutes of Health (NIH), for their detection of IgG antibodies to S. stercoralis. A total of 101 retrospective serum samples, previously submitted for routine S. stercoralis antibody detection using the SciMedx assay, were also evaluated by the InBios and LIPS assays. The qualitative results from each assay were compared using a Venn diagram analysis, to the consensus result among the three assays, and each ELISA was also evaluated using the LIPS assay as the reference standard. By Venn diagram analysis, 65% (66/101) of the samples demonstrated perfect agreement by all three assays. Also, the numbers of samples considered positive or negative by a single method were similar. Compared to the consensus result, the overall percent agreement of the InBios, SciMedx, and LIPS assays were comparable at 87.1%, 84.2%, and 89.1%, respectively. Finally, the two ELISAs performed analogously but demonstrated only moderate agreement (kappa coefficient for the two assays, 0.53) with the LIPS assay. Collectively, while the two commercially available ELISAs perform equivalently, neither should be used independently of clinical evaluation to diagnose strongyloidiasis.