Standard two-tiered testing (STTT) is the recommended algorithm for laboratory diagnosis of Lyme disease (LD). Several limitations are associated with STTT that include low sensitivity in the early stages of disease, as well as technical complexity and subjectivity associated with second-tier immunoblotting; therefore, modified two-tiered testing (MTTT) algorithms that utilize two sequential first-tier tests and eliminate immunoblotting have been evaluated. Recently, a novel MTTT that uses a VlsE chemiluminescence immunoassay followed by a C6 enzyme immunoassay has been proposed. The purpose of this study was to evaluate the performance of the VlsE/C6 MTTT using well-characterized serum samples. Serum samples from the CDC Lyme Serum Repository were tested using three MTTTs, VlsE/C6, whole-cell sonicate (WCS)/C6, and WCS/VlsE, and three STTTs (immunoblotting preceded by three different first-tier assays: VlsE, C6, and WCS). Significant differences were not observed between the results of the MTTTs assessed; however, the VlsE/C6 MTTT resulted in the highest specificity (100%) when other diseases were tested and the lowest sensitivity (75%) for LD samples. Significant differences were present between the results for various MTTTs and STTTs evaluated. Specifically, all MTTTs resulted in higher sensitivities than the STTTs for all LD groups combined and were significantly more accurate (i.e., higher proportion of correct classifications) for this group, with the exception of the WCS/ViraStripe STTT. Additionally, when other diseases were tested, only the results of the VlsE/C6 MTTT differed significantly from those of the WCS/ViraStripe STTT, with the VlsE/C6 MTTT resulting in a 6.2% higher accuracy. Overall, the VlsE/C6 MTTT offers an additional laboratory testing algorithm for LD with equivalent or enhanced performance compared to that of the other MTTTs and STTTs evaluated in this study.

The envelope glycoprotein G of rabies virus in vaccines induces the production of neutralizing antibodies important in the protection against the disease. The measurement of anti-envelope glycoprotein antibodies is a good predictor of the degree of humoral immunity in people during anti-rabies treatment or after vaccination. Several assays exist for the serological determination of antibody protection against rabies virus infection. Antibody neutralization by the rapid fluorescent focus inhibition test (RFFIT) or the fluorescent antibody virus neutralization (FAVN) test is currently the gold standard. Performance of the highly complex RFFIT and FAVN tests, however, requires specialized reference laboratories with expertise with this assay. Although not widely used, ELISA test kits are available and may be an additional option for testing that is more accessible. The aim of the present study was to evaluate available ELISA assays for the determination of anti-rabies antibodies. We compared the Bio-Rad Platelia Rabies II ELISA, DRG Rabies Virus IgG Ab ELISA and Focus Diagnostics Rabies Antibody Detection by ELISA to RFFIT. Bland-Altman plots comparing the Bio-Rad Platelia assay and the Focus Diagnostics assay to RFFIT showed a low degree of variability between the ELISA assays and RFFIT results except in samples with high RFFIT values. The agreement, sensitivity and specificity of Bio-Rad Platelia Rabies II ELISA when compared to RFFIT were 95.1 %, 94.1 % and 95.8 %, respectively. The DRG Rabies assay compared to RFFIT had an agreement of 77.7 %, a sensitivity of 86.7 % and a specificity of 69.4 %. The agreement, sensitivity and specificity of Focus Diagnostics Rabies Detection by ELISA when compared to RFFIT were 82.2 %, 91.7 % and 73.0 %, respectively. Overall, the BioRad Platelia assay showed higher accuracy and specificity than either the DRG or Focus assays. All of these ELISAs, however, measure all antibody types and do not discriminate the neutralizing antibodies as measured by functional assays (RFFIT and FAVN) and cannot be relied upon to predict the neutralizing activity of the sera. The results of this study offer insight into the availability of alternative, less-complex methods to monitor rabies antibody titres in at-risk individuals following vaccination.

Tuberculosis (TB) caused byApproximately nine million new cases of disease and over two million deaths result from tuberculosis (TB) each year (29,56). It is estimated that over one-third of the world's population is infected, with ϳ95% of all cases occurring in developing countries. Global measures attempting to reduce the transmission of TB are currently in place.An essential component of TB control efforts is to identify and treat individuals with active TB disease. The ability to correctly identify individuals with latent TB infection who will progress to active TB disease is vital to this goal (9, 49). Current test procedures are inadequate to accurately detect and identify active TB disease (14,27,30,31,41,44). These shortcomings result in the unnecessary treatment of many individuals who may not need it (3,17,32,45). While the tuberculin skin test (TST) and the QuantiFERON-TB Gold (QFT-G), the traditional methods for latent TB infection screening, rely on the cell-mediated response, the humoral response appears to correlate with the progression of the infection to active TB disease (5,6,11,15,20,21).Many studies have been conducted to evaluate the utility of individual specific Mycobacterium tuberculosis antigens for detecting antibodies in patients with active TB disease (1,7,10,11,20,21,25,26,29,38,39,45,46). Several of these antigens have been developed into commercial assays capable of detecting M. tuberculosis antibodies (4,28,35,53). This study evaluates three commercially available enzyme-linked immunosorbent assays (ELISAs) for their ability to detect immunoglobulin G (IgG) antibodies to M. tuberculosis in patients with active TB disease. MATERIALS AND METHODSHuman sera. The procedures followed were in accordance with the ethical standards established by the University of Utah and are in accordance with the Helsinki Declaration of 1975. This study was approved by the Institutional Review Board of the University of Utah, IRB 17152. All patient samples included in this study were deidentified to meet the Health Information Portability and Accountability Act (HIPAA) patient confidentiality guidelines.Serum samples were stored at Ϫ70°C until testing commenced and were then stored at 2 to 8°C while testing was performed. The whole-blood samples were processed immediately after collection. Samples with discrepant results were tested again by each respective assay to ensure reproducibility. A total of 209 samples were used and divided into four groups.Risk factors that were evaluated for exposure to TB included work in the health care field, laboratory work (especially in mycobacteriology laboratories and specimen processing), immigration from or travel to an country where TB is endemic, and exposure to persons with known active disease. Work in a mycobacteriology laboratory, exposure to a known active case, or immigration from a country where TV is endemic were considered high risk for exposure. Work in a health care field was considered moderate risk.Group I serum samples consisted of 88 samples from h...

ABSTRACTAnti-Mycobacterium tuberculosisIgG antibodies may aid in the diagnosis of activeM. tuberculosisdisease. We studied whether anti-M. tuberculosisIgG antibodies are elevated in activeM. tuberculosisdisease and assessed factors contributing to false-positive and -negative results. A retrospective study of 2,150 individuals tested by the QuantiFERON-TB Gold In-Tube (QFT-GIT) assay was conducted at the University of Utah, ARUP Laboratories, November 2008 to December 2010. All samples were tested with the InBios Active TbDetectantituberculosis (anti-TB) IgG antibody assay. Of 1,044 patients with a positive QFT-GIT, 59 (5.7%) were positive forM. tuberculosisantibodies. Fourteen of 1,106 (1.3%) with a negative or indeterminate QFT-GIT were positive forM. tuberculosisantibodies.M. tuberculosisantibody tests were positive in 61.5% with confirmed activeM. tuberculosisdisease and other mycobacterial infections. Over half of the false-negativeM. tuberculosisantibody tests occurred in patients ≥90 years of age. False positives were seen in 12.9% of autoimmune patients. The odds ratio of being positive by the QFT-GIT and the InBios TB IgG assay increased with confirmedM. tuberculosisdisease or highly suspectedM. tuberculosisdisease and was 86.7 (95% confidence interval [CI], 34.4 to 218.5) in these two groups compared to patients negative by both tests. Although anti-M. tuberculosisantibodies can be detected in patients with activeM. tuberculosisdisease, caution should be used with patients where immunoglobulin levels may be decreased or patients with autoantibodies.

In the present study, two immunoglobulin G (IgG) immunoblot assays and one IgG Western blot assay were compared to the rapid plasma reagin test (RPR), the fluorescent treponemal antibody absorption test (FTA-ABS), and the Treponema pallidum particle agglutination assay (TP-PA). The agreement levels of the Viramed, Virotech, and MarDx assays were 97.0%, 96.4%, and 99.4%, and the agreements of samples inconclusive by FTA-ABS and resolved by TP-PA were 91.7%, 83.3%, and 69.4%, respectively.Syphilis, a disease caused by Treponema pallidum, is transmitted congenitally or through sexual intercourse (8-9). Nontreponema-based tests such as the rapid plasma reagin test (RPR) are used to detect syphilis infection (6, 9-10). These tests may produce false-positive results in pregnant women and patients with infections (3, 5-6, 9, 11). An algorithm has been developed for the serological diagnosis of syphilis which includes a non-treponema-based screening test and a treponemabased confirmatory assay (1-2, 7, 11). Traditional confirmatory assays include the fluorescent treponemal antibody absorption test (FTA-ABS) and the T. pallidum particle agglutination assay (TP-PA) (9).Western blot-based assays to detect immunoglobulin G (IgG) antibodies may prove useful, especially in cases where the FTA-ABS is inconclusive. In the present study, results of two immunoblot assays and one Western blot assay were compared to FTA-ABS/TP-PA and RPR results, as well as to each other.Human sera. A total of 200 human serum samples sent to Associated Regional and University Pathologists (ARUP) laboratories for syphilis testing were collected. Procedures were followed in accordance with the ethical standards established by the University of Utah in accordance with the Helsinki Declaration of 1975. All patient samples were deidentified according to the University of Utah Institutional Review Board protocol (no. 7275) to meet the Health Information Portability and Accountability Act guidelines. Specimens were stored at Ϫ20°C until testing and then stored at 2 to 8°C.Non-treponema-based testing. All 200 samples were tested by RPR according to the manufacturer's protocol (Arlington Scientific, Inc., Springville, UT).Treponema-based testing. One hundred forty-two samples were tested by FTA-ABS (Inverness Medical, Waltham, MA), and 32 inconclusive samples were further tested by TP-PA (Fujirebio, Malvern, PA). Both assays were performed according to the manufacturers' protocols. The 32 inconclusive FTA-ABS samples were included to reflect the high percentage of inconclusive FTA-ABS samples sent to our reference laboratory from primary screening laboratories.Syphilis blot testing. All 200 samples were tested using two immunoblot assays and one Western blot assay, the Treponema ViraBlot test kit IgG (Viralab Inc., Oceanside, CA), the Treponema pallidum IgG line immunoblot (Genzyme Virotech GmbH, Rüsselsheim, Germany), and the T. pallidum IgG Marblot strip test system (MarDx Diagnostics, Inc., Carlsbad, CA). Each assay was performed according to the manufac...