Publications by authors named "Michael T Pyne"

18 Publications

  • Page 1 of 1

Efficient and effective single-step screening of individual samples for SARS-CoV-2 RNA using multi-dimensional pooling and Bayesian inference.

J R Soc Interface 2021 06 16;18(179):20210155. Epub 2021 Jun 16.

Department of Chemistry, University of Utah, Salt Lake City, UT, USA.

Rapid and widespread implementation of infectious disease surveillance is a critical component in the response to novel health threats. Molecular assays are the preferred method to detect a broad range of viral pathogens with high sensitivity and specificity. The implementation of molecular assay testing in a rapidly evolving public health emergency, such as the ongoing COVID-19 pandemic, can be hindered by resource availability or technical constraints. We present a screening strategy that is easily scaled up to support a sustained large volume of testing over long periods of time. This non-adaptive pooled-sample screening protocol employs Bayesian inference to yield a reportable outcome for each individual sample in a single testing step (no confirmation of positive results required). The proposed method is validated using clinical specimens tested using a real-time reverse transcription polymerase chain reaction test for SARS-CoV-2. This screening protocol has substantial advantages for its implementation, including higher sample throughput, faster time to results, no need to retrieve previously screened samples from storage to undergo retesting, and excellent performance of the algorithm's sensitivity and specificity compared with the individual test's metrics.
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http://dx.doi.org/10.1098/rsif.2021.0155DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8205536PMC
June 2021

In Search of Covariates of HIV-1 Subtype B Spread in the United States-A Cautionary Tale of Large-Scale Bayesian Phylogeography.

Viruses 2020 02 5;12(2). Epub 2020 Feb 5.

Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, KU Leuven, 3000 Leuven, Belgium.

Infections with HIV-1 group M subtype B viruses account for the majority of the HIV epidemic in the Western world. Phylogeographic studies have placed the introduction of subtype B in the United States in New York around 1970, where it grew into a major source of spread. Currently, it is estimated that over one million people are living with HIV in the US and that most are infected with subtype B variants. Here, we aim to identify the drivers of HIV-1 subtype B dispersal in the United States by analyzing a collection of 23,588 ol sequences, collected for drug resistance testing from 45 states during 2004-2011. To this end, we introduce a workflow to reduce this large collection of data to more computationally-manageable sample sizes and apply the BEAST framework to test which covariates associate with the spread of HIV-1 across state borders. Our results show that we are able to consistently identify certain predictors of spread under reasonable run times across datasets of up to 10,000 sequences. However, the general lack of phylogenetic structure and the high uncertainty associated with HIV trees make it difficult to interpret the epidemiological relevance of the drivers of spread we are able to identify. While the workflow we present here could be applied to other virus datasets of a similar scale, the characteristic star-like shape of HIV-1 phylogenies poses a serious obstacle to reconstructing a detailed evolutionary and spatial history for HIV-1 subtype B in the US.
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http://dx.doi.org/10.3390/v12020182DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7077180PMC
February 2020

Clinical Performance of SurePath™ Preservative Compared to PreservCyt® with Cobas® and Hybrid Capture® 2 HPV Tests in a Colposcopy Population.

Clin Lab 2019 Mar;65(3)

Background: Until recently, no HPV test had been US FDA-approved for SurePath preservative. Clinical performance remains incompletely understood. The clinical performances of the Cobas HPV Test (Cobas) and Hybrid Capture 2 High-Risk HPV DNA Test (HC2) with PreservCyt and SurePath preservatives were compared.

Methods: Cervical cytology samples were collected in both preservatives in random order from women age 21+ (n = 244) referred for colposcopy. Before cytology processing and pelleting, SurePath samples were tested by the Cobas test with and without buffered SDS heat pretreatment. SurePath pellets were tested by the HC2 test and by the Cobas test (with pretreatment). Performance characteristics were calculated in relation to cases of cervical in-traepithelial neoplasia grade 2 or higher (CIN2+) as the clinical target outcome. All HPV-positive samples were also genotyped with the Linear Array test.

Results: CIN2+ was detected in 42 patients (17.2%). For both HPV tests, there was a trend towards higher positivity and sensitivity for SurePath compared to PreservCyt preservative. The Cobas test had higher sensitivity than HC2 and the HC2 test had higher specificity than Cobas. Pretreated SurePath samples produced results similar to untreated ones, despite a two-fold dilution during pretreatment [sensitivity %: 95.1 (82.2 - 99.2) vs. 94.3 (79.5 - 99.0); specificity %: 33.0 (26.6 - 40.1) vs. 33.0 (26.4 - 40.3)].

Conclusions: There was good agreement between the preservatives and HPV tests in detecting HPV and between the Cobas and Linear Array tests for genotyping HR-HPV. These trends were not statistically significant due to the limited number of CIN2+ cases. However, these data may help in evaluations of preservative selection for colposcopy samples. Pre-treatment for Cobas testing eliminated invalid results due to clots. The Cobas test has been FDA-approved for use with heat pretreated SurePath samples.
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http://dx.doi.org/10.7754/Clin.Lab.2018.180734DOI Listing
March 2019

Cervical Cytology Specimen Stability in Surepath Preservative and Analytical Sensitivity for HPV Testing with the cobas and Hybrid Capture 2 Tests.

PLoS One 2016 23;11(2):e0149611. Epub 2016 Feb 23.

ARUP Institute for Clinical and Experimental Pathology, Salt Lake City, UT, United States of America.

None of the commercial HPV tests are U.S. FDA-approved for testing of cervical cytology specimens in SurePath preservative. Still, ~30% of HPV testing is performed on specimens in this formalin-containing preservative. Formalin-induced DNA fragmentation and cross-linking may interfere with HPV detection. We evaluated analytical sensitivity and specimen stability of the cobas 4800 HPV (Roche) and Hybrid Capture 2 HPV (HC2, Qiagen) tests with residual cervical cytology samples in SurePath preservative available within 1 week of collection. Cobas testing was performed with and without heating samples at 120°C for 20 min diluted 1:1 in an alkaline environment (pretreatment) to revert DNA crosslinking. Stability was tested after 2 weeks of storage at ambient temperature followed by ≤10 weeks at 4°C. Analytical sensitivity and positivity rates (HC2, 18%; cobas pretreated, 46%; cobas untreated, 47%) were greater for cobas than HC2 (n = 682). After 6 weeks of storage, mean HC2 ratios were lower (mean 0.9, SD 6.3) but high variability limited statistical power to detect trends. Cobas threshold cycles (Ct's) increased in untreated (mean 2.1) but not pretreated samples (mean 0.3; n = 110; p≤0.0001). Overall, cobas had greater analytical sensitivity for samples in SurePath preservative. Although pretreatment introduced a manual sample transfer step and 30 min of incubation times, it improved stability without negatively affecting analytical sensitivity. While awaiting results of large trials to evaluate the clinical performance of cobas, the addition of the pretreatment step may improve the detection of HPV, especially after prolonged sample storage.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0149611PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4764333PMC
July 2016

High-risk HPV detection and genotyping by APTIMA HPV using cervical samples.

J Virol Methods 2015 Sep 5;221:95-9. Epub 2015 May 5.

ARUP Institute for Clinical and Experimental Pathology, 500 Chipeta Way, Salt Lake City, UT 84108, USA; ARUP Laboratories, 500 Chipeta Way, Salt Lake City, UT 84108, USA. Electronic address:

High-risk human papillomavirus (HPV) detection and genotyping is critical for cervical cancer screening. Testing of 967 cervical cytology specimens in PreservCyt preservative revealed similar positivity rates for HC2 (13.8%) and APTIMA HPV (AHPV) tests (13.5%, p=0.89) and high overall agreement (94.6%, κ=0.77). A trend towards higher HPV16 positivity rates by the Cobas HPV test (23.0%, 26/113) compared to the AHPV genotyping assay (19.5%, 22/113; p=0.125) was noted. No cross-contamination was detected with AHPV in a challenge experiment.
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http://dx.doi.org/10.1016/j.jviromet.2015.04.030DOI Listing
September 2015

HCV RNA measurement in samples with diverse genotypes using versions 1 and 2 of the Roche COBAS® AmpliPrep/COBAS® TaqMan® HCV test.

J Clin Virol 2015 Apr 7;65:54-7. Epub 2015 Feb 7.

ARUP Institute for Clinical and Experimental Pathology(®), 500 Chipeta Way, Salt Lake City, UT 84108, USA; University of Utah Department of Pathology, 15 N Medical Drive East, Salt Lake City, UT 84112, USA.

Background: Accurate HCV RNA measurement is required for monitoring treatment. Underquantification has been reported with some genotypes, particularly genotype 4, using version 1 of the Roche COBAS(®) AmpliPrep/COBAS(®) TaqMan(®) HCV Test.

Objectives: Compare the viral loads of clinical specimens representing diverse genotypes from across the United States using versions 1 (V1) and 2 (V2) of the Roche COBAS(®) AmpliPrep/COBAS(®) TaqMan(®) HCV Test. Assess the frequency of nt145 and nt165 variants in the 5' UTR associated with detection failures and underquantification in a large clinical sample database.

Study Design: Three hundred archived clinical samples were measured using V1 and V2. Bland-Altman analysis was performed on log-transformed results and compared by genotype. The frequencies of nt145 and nt165 variants from 15,817 sequences were calculated.

Results: On average, V2 results were 0.16 logIU/mL lower than V1 results. The average genotype 4 sample was 0.08 logIU/mL higher in V1 than V2. The largest individual sample differences were -2.10 (genotype 2b) and 1.57 (genotype 2a) logIU/mL. For genotype 4 samples, the greatest underquantification by V1 was 1.46 logIU/mL. There were 13 (0.082%) variants at nt145 and 24 variants at nt165 (0.152%), including one sequence with variants at both positions (0.0063%).

Conclusions: Genotype 4 samples from the U.S. are rarely underquantified and not disproportionately so compared to other genotypes using the COBAS(®) AmpliPrep/COBAS(®) TaqMan(®) HCV Tests. Variants at the nt145 and nt165 positions are uncommon in the U.S. and double variants are exceedingly rare. Underquantification of HCV samples with the V1 assay is likely a very rare occurrence in U.S.
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http://dx.doi.org/10.1016/j.jcv.2015.02.002DOI Listing
April 2015

Testing and genotyping of high-risk human papillomavirus by the cobas HPV Test and the Hybrid Capture 2 high-risk HPV DNA test using cervical and vaginal samples.

J Clin Microbiol 2014 May 19;52(5):1720-3. Epub 2014 Feb 19.

ARUP Institute for Clinical and Experimental Pathology, Salt Lake City, Utah, USA.

The accurate detection and typing of high-risk human papillomavirus (HPV) are critical for cervical cancer screening. The Hybrid Capture 2 (hc2) and cobas HPV tests showed high agreement for cervical samples (94.4%, κ=0.72, n=693) and moderate agreement for vaginal samples (κ=0.62, n=108). The HPV16 and HPV18 results were highly consistent between the cobas and Linear Array tests (κ≥0.96, n=197). Three hc2-negative vaginal samples were repeatedly invalid by the cobas test due to β-globin control failures, highlighting amplification control benefits. No cross-contamination was detected in a challenge experiment.
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http://dx.doi.org/10.1128/JCM.03308-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3993637PMC
May 2014

Evaluation of the Roche COBAS AmpliPrep/COBAS TaqMan HCV Test.

Diagn Microbiol Infect Dis 2013 Sep 24;77(1):25-30. Epub 2013 Jun 24.

ARUP Institute for Clinical and Experimental Pathology®, Salt Lake City, UT 84108, USA.

The performance of the Roche COBAS AmpliPrep/COBAS TaqMan HCV Test (CAP/CTM) was evaluated. The limit of detection was 9.5 IU/mL using the 3rd International Standard. Serial dilutions of each genotype demonstrated good reproducibility and linearity. Correlation with samples previously tested with the Roche Analyte Specific Reagent (ASR) was very good, with the CAP/CTM assay measuring 0.24 log IU/mL higher on average than ASR. Genotype inclusivity evaluated in the CAP/CTM, ASR, and Siemens Versant HCV RNA 3.0 Assay (bDNA) assay using a commercially available panel showed higher measurements than ASR or bDNA. The differences in observed CAP/CTM and ASR results for genotype 3 patient samples were significantly different (P < 0.05) from those for both genotype 1 and 2 samples. Common inhibitory substances had no more than a 0.25 log IU/mL affect. Overall, the automated CAP/CTM assay exhibits excellent sensitivity, reproducibility, and dynamic range. Its performance is compatible with its use in guiding therapy using direct acting antivirals such as boceprevir and telaprevir.
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http://dx.doi.org/10.1016/j.diagmicrobio.2013.05.012DOI Listing
September 2013

Large-scale analysis of the prevalence and geographic distribution of HIV-1 non-B variants in the United States.

J Clin Microbiol 2013 Aug 12;51(8):2662-9. Epub 2013 Jun 12.

ARUP Institute for Clinical and Experimental Pathology, Salt Lake City, Utah, USA.

The genetic diversity of human immunodeficiency virus type 1 (HIV-1) has significant implications for diagnosis, vaccine development, and clinical management of patients. Although HIV-1 subtype B is predominant in the United States, factors such as global travel, immigration, and military deployment have the potential to increase the proportion of non-subtype B infections. Limited data are available on the prevalence and distribution of non-B HIV-1 strains in the United States. We sought to retrospectively examine the prevalence, geographic distribution, diversity, and temporal trends of HIV-1 non-B infections in samples obtained by ARUP Laboratories, a national reference laboratory, from all regions of the United States. HIV-1 pol sequences from 24,386 specimens collected from 46 states between 2004 and September 2011 for drug resistance genotyping were analyzed using the REGA HIV-1 Subtyping Tool, version 2.0. Sequences refractory to subtype determination or reported as non-subtype B by this tool were analyzed by PHYLIP version 3.5 and Simplot version 3.5.1. Non-subtype B strains accounted for 3.27% (798/24,386) of specimens. The 798 non-B specimens were received from 37 states and included 5 subtypes, 23 different circulating recombinant forms (CRFs), and 39 unique recombinant forms (URFs). The non-subtype B prevalence varied from 0% in 2004 (0/54) to 4.12% in 2011 (201/4,884). This large-scale analysis reveals that the diversity of HIV-1 in the United States is high, with multiple subtypes, CRFs, and URFs circulating. Moreover, the geographic distribution of non-B variants is widespread. Data from HIV-1 drug resistance testing have the potential to significantly enhance the surveillance of HIV-1 variants in the United States.
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http://dx.doi.org/10.1128/JCM.00880-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3719628PMC
August 2013

Sequencing-based genotyping of mixed human papillomavirus infections by use of RipSeq software.

J Clin Microbiol 2013 Apr 30;51(4):1278-80. Epub 2013 Jan 30.

ARUP Institute for Clinical and Experimental Pathology, Salt Lake City, Utah, USA.

Sequencing-based pathogen identification directly from clinical specimens requires time-consuming interpretation, especially with mixed chromatograms when multiple microorganisms are detected. We assessed RipSeq Mixed software for human papillomavirus (HPV) genotyping by comparison to the linear array HPV genotyping assay. RipSeq Mixed provided rapid, sequencing-based HPV typing for single-type infections and coinfections with 2 types.
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http://dx.doi.org/10.1128/JCM.00091-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3666769PMC
April 2013

A retrospective review of UroVysion fish interpretations over 8.6 years: a major shift in the patient test population.

Diagn Cytopathol 2013 May 4;41(5):437-47. Epub 2012 Aug 4.

ARUP Institute for Clinical and Experimental Pathology®, ARUP Laboratories, Inc, Salt Lake City, Utah, USA.

UroVysion FISH detects chromosomal aberrations associated with urothelial carcinoma. In our laboratory, UroVysion FISH was initially evaluated manually with a change to image-aided interpretation using the BioView Duet imaging system. This retrospective study examined diagnostic findings over an 8.6 year period, with 1,869 manual interpretations over 4.8 years and 3,936 image-aided interpretations over 3.8 years. Although the initial goal was to evaluate possible impacts of the imaging system on diagnostic interpretations, the most important finding was that the demographics of the test population changed significantly. Female specimens increased incrementally from an average of 29% compared to 43% of the samples during periods of manual interpretation versus image-aided interpretation, respectively. The shift may reflect a gradual increase in the percentage of low-risk hematuria patients being evaluated for initial diagnosis of bladder cancer, rather than bladder cancer recurrence. Interpretation rates, evaluated separately for males and females, changed significantly over the test period. Male interpretation results were negative (75.1 vs. 67%), positive (18.6 vs. 14.6%), unsatisfactory (5.0 vs. 16.9%), and equivocal (1.4 vs. 1.5%) during periods of manual versus image-aided interpretation, respectively (Fisher Exact Test P-value = <0.0001). For females, results were negative (86.1 vs. 79.3%), positive (9.2 vs. 11.1%), unsatisfactory (2.8 vs. 8.9%), and equivocal (1.8 vs. 0.7%) over the same periods (Fisher Exact Test P-value = <0.0001). Logistic regression analysis identified the change in test population as the variable with the greatest impact on observed interpretation rate changes.
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http://dx.doi.org/10.1002/dc.22881DOI Listing
May 2013

Large-scale comparison of Roche Cobas AmpliPrep/Cobas TaqMan and Abbott RealTime HIV assays.

J Virol Methods 2012 Sep 17;184(1-2):106-8. Epub 2012 May 17.

ARUP Institute for Clinical and Experimental Pathology®, 500 Chipeta Way, Salt Lake City, UT 84108, United States.

Significant underquantitation of HIV RNA has been reported with the Roche Cobas AmpliPrep/Cobas TaqMan HIV Test (Version 1) compared to other assays. However, these studies have generally involved limited numbers of samples from select patient populations or analysis of samples that were undetectable in the TaqMan assay. Random plasma samples submitted from throughout the United States for HIV RNA quantitation (n=1263) were compared in the Roche TaqMan and Abbott RealTime assays. Twenty-four samples (1.9%) were discrepant, with a maximum difference between the two assays of 1.9logcopies/mL. These data indicate that both tests may be susceptible to underquantitation, but the incidence is low in this large cohort of samples from across the United States.
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http://dx.doi.org/10.1016/j.jviromet.2012.05.007DOI Listing
September 2012

Comparison of three Roche hepatitis B virus viral load assay formats.

J Clin Microbiol 2012 Jul 25;50(7):2337-42. Epub 2012 Apr 25.

ARUP Institute for Clinical and Experimental Pathology, Salt Lake City, Utah, USA.

Two FDA-approved (in vitro diagnostic [IVD]) hepatitis B virus (HBV) viral load assays, the manual Cobas TaqMan HBV Test for use with the High Pure System (HP) and the automated Cobas AmpliPrep/Cobas TaqMan HBV Test v2.0 (CAP/CTM), were compared to a modified (not FDA-approved) version of the HP assay by automating the DNA extraction using the Total Nucleic Acid Isolation (TNAI) kit on the Cobas AmpliPrep. On average, CAP/CTM measurements were 0.08 log IU/ml higher than HP results (n = 206), and TNAI results were 0.17 log IU/ml higher than HP results (n = 166). The limit of detection (LOD), as determined by probit analysis using dilutions of the 2nd HBV international standard, was 10.2 IU/ml for CAP/CTM. The data sets for HP and TNAI were insufficient for probit analysis; however, there was 100% detection at ≥ 5 or ≥ 10 IU/ml for TNAI and HP, respectively. Linearity was demonstrated between 60 and 2,000,000 IU/ml, with slopes between 0.95 and 0.99 and R(2) values of >0.99 for all assays. Total precision (log percent coefficient of variance [CV]) was between 0.8% and 2.1% at 4.3 log IU/ml and between 1.4% and 4.9% at 2.3 log IU/ml. Correlation of samples, reproducibility, linearity, and LOD were acceptable and similar in all assays. The CAP/CTM assay and, to a lesser extent, the TNAI assay reduced hands-on time due to automation. There were no instances of contamination detected in negative samples during the course of the study, despite testing several samples up to 9.6 log IU/ml. The incidence of false-positive negative controls in HP and CAP/CTM clinical testing was <0.5% over 6 to 7 months of testing.
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http://dx.doi.org/10.1128/JCM.00746-12DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3405597PMC
July 2012

Performance of the COBAS® AmpliPrep/COBAS TaqMan® automated system for hepatitis C virus (HCV) quantification in a multi-center comparison.

J Clin Virol 2011 Feb 9;50(2):100-3. Epub 2010 Dec 9.

University of Iowa, Roy J and Lucille A. Carver College of Medicine, Department of Pathology, 200 Hawkins Drive, Iowa City, IA 52242, USA.

Background: Quantitative HCV RNA testing is considered standard of care for monitoring during treatment of patients infected with HCV. The COBAS(®) AmpliPrep/COBAS(®) TaqMan(®) HCV Test fully automates specimen processing and reaction assembly for HCV viral load testing using reverse transcription and real-time PCR amplification.

Objectives: The performance of the COBAS(®) AmpliPrep/COBAS(®) TaqMan(®) HCV Test was evaluated in a multi-center study.

Study Design: Typical plasma based specimens were tested for accuracy, analytic range of measurement, reproducibility and genotype specific quantitation.

Results: Linear regression analysis of the quantitative results demonstrated a linear range of detection from 50 to 5 million (1.7-6.7 log(10))IU/mL and a coefficient of determination (R(2)) of 0.9948. The precision of the assay was highly reproducible within and between runs and among laboratories with coefficients of variance (CV) ranging from 6.7% to 40.0% across the seven laboratories. A representative sample for each of the six major HCV genotypes demonstrated reproducible quantitation between the seven laboratories.

Conclusions: The COBAS(®) AmpliPrep/COBAS(®) TaqMan(®) HCV Test is a reliable and sensitive assay for HCV RNA quantitation.
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http://dx.doi.org/10.1016/j.jcv.2010.10.020DOI Listing
February 2011

Evaluation of the Roche Cobas AmpliPrep/Cobas TaqMan HIV-1 test and identification of rare polymorphisms potentially affecting assay performance.

J Clin Microbiol 2010 Aug 23;48(8):2852-8. Epub 2010 Jun 23.

ARUP Institute for Clinical and Experimental Pathology, 500 Chipeta Way, Salt Lake City, UT 84108, USA.

We evaluated the FDA-approved Roche Cobas AmpliPrep/Cobas TaqMan (CAP/CTM) HIV-1 viral load assay for sensitivity, reproducibility, linearity, HIV-1 subtype detection, and correlation to the Roche Amplicor HIV-1 monitor test, version 1.5 (Amplicor). The limit of detection calculated by probit analysis was 23.8 copies/ml using the 2nd International WHO Standard and 30.8 copies/ml using Viral Quality Assurance (VQA) standard material. Serial dilutions of six patient samples were used to determine inter- and intra-assay reproducibility and linearity, which were very good (<8% coefficient of variation [CV]; between approximately 1.7 and 7.0 log(10) copies/ml). Subtype detection was evaluated in the CAP/CTM, Amplicor, and Bayer Versant HIV-1 bDNA 3.0 (Versant) assays using a commercially available panel. Versant averaged 0.829 log(10) copies/ml lower than CAP/CTM and Amplicor averaged 0.427 log(10) copies/ml lower than CAP/CTM for the subtype panel. Correlation with samples previously tested by Amplicor was excellent (R(2) = 0.884; average difference [Amplicor value minus CAP/CTM value], 0.008 log(10) copies/ml). Of the 305 HIV samples tested, 7 samples generated CAP/CTM titers between 1.0 and 2.75 log(10) copies/ml lower than those for Amplicor. Three of these samples revealed primer and probe mismatches that could account for the discrepancies. Otherwise, the CAP/CTM assay exhibits excellent sensitivity, dynamic range, reproducibility, and correlation with Amplicor in an automated format.
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http://dx.doi.org/10.1128/JCM.00776-10DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2916612PMC
August 2010

Evaluation of the Abbott investigational use only RealTime hepatitis C virus (HCV) assay and comparison to the Roche TaqMan HCV analyte-specific reagent assay.

J Clin Microbiol 2009 Sep 22;47(9):2872-8. Epub 2009 Jul 22.

ARUP Institute for Clinical and Experimental Pathology, 500 Chipeta Way, Salt Lake City, UT 84108, USA.

The accurate and sensitive measurement of hepatitis C virus (HCV) RNA is essential for the clinical management and treatment of infected patients and as a research tool for studying the biology of HCV infection. We evaluated the linearity, reproducibility, precision, limit of detection, and concordance of viral genotype quantitation of the Abbott investigational use only RealTime HCV (RealTime) assay using the Abbott m2000 platform and compared the results to those of the Roche TaqMan Analyte-Specific Reagent (TaqMan) and Bayer Versant HCV bDNA 3.0 assay. Comparison of 216 samples analyzed by RealTime and TaqMan assays produced the following Deming regression equation: RealTime = 0.940 (TaqMan) + 0.175 log(10) HCV RNA IU/ml. The average difference between the assays was 0.143 log(10) RNA IU/ml and was consistent across RealTime's dynamic range of nearly 7 log(10) HCV RNA IU/ml. There was no significant difference between genotypes among these samples. The limit of detection using eight replicates of the World Health Organization HCV standard was determined to be 7.74 HCV RNA IU/ml by probit analysis. Replicate measurements of commercial genotype panels were significantly higher than TaqMan measurements for most samples and showed that the RealTime assay is able to detect all genotypes with no bias. Additionally, we showed that the amplicon generated by the widely used Roche COBAS Amplicor Hepatitis C Virus Test, version 2.0, can act as a template in the RealTime assay, but potential cross-contamination could be mitigated by treatment with uracil-N-glycosylase. In conclusion, the RealTime assay accurately measured HCV viral loads over a broad dynamic range, with no significant genotype bias.
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http://dx.doi.org/10.1128/JCM.02329-08DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2738061PMC
September 2009

Evaluation of the Abbott investigational use only realtime HIV-1 assay and comparison to the Roche Amplicor HIV-1 monitor test, version 1.5.

J Mol Diagn 2009 Jul 21;11(4):347-54. Epub 2009 May 21.

ARUP Institute for Clinical and Experimental Pathology, Salt Lake City, UT 84108, USA.

Abbott Molecular's m2000 system and RealTime HIV-1 assay (RealTime) were evaluated for sensitivity, reproducibility, linearity, ability to detect diverse HIV-1 subtypes/groups, and correlation to the Roche AMPLICOR HIV-1 MONITOR Test, Version 1.5 (Amplicor). The limit of detection was determined using the second International World Health Organization Standard and Viral Quality Assurance standard material. Serial dilutions of four patient samples were used to determine inter- and intra-assay reproducibility and linearity. Samples representing HIV-1 groups M, N, and O were evaluated in the RealTime, Amplicor, and Siemens Versant HIV-1 branched chain DNA 3.0 (Versant) assays. Archived Amplicor-tested samples were tested with the 1 ml, 0.5 ml, and 0.6 ml versions of the RealTime assay. Probit analysis predicts a limit of detection of 21.94 IU/ml using the World Health Organization Standard and 26.54 copies/ml using Viral Quality Assurance material with the 1 ml assay. Linearity and reproducibility were very good between approximately 1.60 to 6.0 log(10) copies/ml. All three assays produced similar measurements for all Group M subtypes tested; the RealTime assay was the only assay that detected all three Group O samples tested. Correlation with the Amplicor assay was good, although the RealTime assay measured between 0.342 and 0.716 log(10) copies/ml lower on average, depending on the input volume. The automated RealTime assay exhibits excellent sensitivity, dynamic range, reproducibility, and group/subtype detection, albeit with consistently lower values than Amplicor.
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http://dx.doi.org/10.2353/jmoldx.2009.080166DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2710712PMC
July 2009

A biochemical analysis demonstrates that the BRCA1 intronic variant IVS10-2A--> C is a mutation.

J Hum Genet 2003 ;48(8):399-403

Technology Development, Myraid Genetic Laboratories, Salt Lake City, UT 84108, USA.

Sequence analysis of cDNA from an asymptomatic patient belonging to a high-risk breast cancer family carrying the genetic variant BRCA1 IVS10-2A-->C revealed that functional BRCA1 mRNA was derived from only one of the patient's chromosomes. The other chromosome produced an aberrant RNA splicing transcript that deleted exon 11. Analysis of the patient's genomic DNA demonstrated that the chromosome producing the non-functional mRNA carried the genotype BRCA1 IVS10-2A-->C. This transversion disrupts a highly conserved base in the consensus splice acceptor motif. These results support the conclusion that BRCA1 IVS10-2A-->C is a mutation that confers predisposition to breast and ovarian cancer.
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http://dx.doi.org/10.1007/s10038-003-0044-0DOI Listing
November 2003