Browsing by Subject "Quality Control"
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Item Open Access Automated quality control in nuclear medicine using the structured noise index.(Journal of applied clinical medical physics, 2020-04) Nelson, Jeffrey S; Samei, EhsanPurpose
Daily flood-field uniformity evaluation serves as the central element of nuclear medicine (NM) quality control (QC) programs. Uniformity images are traditionally analyzed using pixel value-based metrics, that is, integral uniformity (IU), which often fail to capture subtle structure and patterns caused by changes in gamma camera performance, requiring visual inspections which are subjective and time demanding. The goal of this project was to implement an advanced QC metrology for NM to effectively identify nonuniformity issues, and report issues in a timely manner for efficient correction prior to clinical use. The project involved the implementation of the program over a 2-year period at a multisite major medical institution.Methods
Using a previously developed quantitative uniformity analysis metric, the structured noise index (SNI) [Nelson et al. (2014), \textit{J Nucl Med.}, \textbf{55}:169-174], an automated QC process was developed to analyze, archive, and report on daily NM QC uniformity images. Clinical implementation of the newly developed program ran in parallel with the manufacturer's reported IU-based QC program. The effectiveness of the SNI program was evaluated over a 21-month period using sensitivity and coefficient of variation statistics.Results
A total of 7365 uniformity QC images were analyzed. Lower level SNI alerts were generated in 12.5% of images and upper level alerts in 1.7%. Intervention due to image quality issues occurred on 26 instances; the SNI metric identified 24, while the IU metric identified eight. The SNI metric reported five upper level alerts where no clinical engineering intervention was deemed necessary.Conclusion
An SNI-based QC program provides a robust quantification of the performance of gamma camera uniformity. It operates seamlessly across a fleet of multiple camera models and, additionally, provides effective workflow among the clinical staff. The reliability of this process could eliminate the need for visual inspection of each image, saving valuable time, while enabling quantitative analysis of inter- and intrasystem performance.Item Open Access Citizen Science as a New Tool in Dog Cognition Research.(PLoS One, 2015) Stewart, L; MacLean, EL; Ivy, D; Woods, V; Cohen, E; Rodriguez, K; McIntyre, M; Mukherjee, S; Call, J; Kaminski, J; Miklósi, Á; Wrangham, RW; Hare, BFamily dogs and dog owners offer a potentially powerful way to conduct citizen science to answer questions about animal behavior that are difficult to answer with more conventional approaches. Here we evaluate the quality of the first data on dog cognition collected by citizen scientists using the Dognition.com website. We conducted analyses to understand if data generated by over 500 citizen scientists replicates internally and in comparison to previously published findings. Half of participants participated for free while the other half paid for access. The website provided each participant a temperament questionnaire and instructions on how to conduct a series of ten cognitive tests. Participation required internet access, a dog and some common household items. Participants could record their responses on any PC, tablet or smartphone from anywhere in the world and data were retained on servers. Results from citizen scientists and their dogs replicated a number of previously described phenomena from conventional lab-based research. There was little evidence that citizen scientists manipulated their results. To illustrate the potential uses of relatively large samples of citizen science data, we then used factor analysis to examine individual differences across the cognitive tasks. The data were best explained by multiple factors in support of the hypothesis that nonhumans, including dogs, can evolve multiple cognitive domains that vary independently. This analysis suggests that in the future, citizen scientists will generate useful datasets that test hypotheses and answer questions as a complement to conventional laboratory techniques used to study dog psychology.Item Open Access Development and implementation of a proficiency testing program for Luminex bead-based cytokine assays.(Journal of Immunological Methods, 2014-07) Lynch, Heather E; Sanchez, Ana M; D'Souza, M Patricia; Rountree, Wes; Denny, Thomas N; Kalos, Michael; Sempowski, Gregory DLuminex bead array assays are widely used for rapid biomarker quantification due to the ability to measure up to 100 unique analytes in a single well of a 96-well plate. There has been, however, no comprehensive analysis of variables impacting assay performance, nor development of a standardized proficiency testing program for laboratories performing these assays. To meet this need, the NIH/NIAID and the Cancer Immunotherapy Consortium of the Cancer Research Institute collaborated to develop and implement a Luminex assay proficiency testing program as part of the NIH/NIAID-sponsored External Quality Assurance Program Oversight Laboratory (EQAPOL) at Duke University. The program currently monitors 25 domestic and international sites with two external proficiency panels per year. Each panel includes a de-identified commercial Luminex assay kit with standards to quantify human IFNγ, TNFα, IL-6, IL-10 and IL-2, and a series of recombinant cytokine-spiked human serum samples. All aspects of panel development, testing and shipping are performed under GCLP by EQAPOL support teams. Following development testing, a comprehensive site proficiency scoring system comprised of timeliness, protocol adherence, accuracy and precision was implemented. The overall mean proficiency score across three rounds of testing has remained stable (EP3: 76%, EP4: 75%, EP5: 77%); however, a more detailed analysis of site reported results indicates a significant improvement of intra- (within) and inter- (between) site variation, suggesting that training and remediation for poor performing sites may be having a positive impact on proficiency. Through continued proficiency testing, identification of variables affecting Luminex assay outcomes will strengthen efforts to bring standardization to the field.Item Open Access Development of a contemporary globally diverse HIV viral panel by the EQAPOL program.(J Immunol Methods, 2014-07) Sanchez, Ana M; DeMarco, C Todd; Hora, Bhavna; Keinonen, Sarah; Chen, Yue; Brinkley, Christie; Stone, Mars; Tobler, Leslie; Keating, Sheila; Schito, Marco; Busch, Michael P; Gao, Feng; Denny, Thomas NThe significant diversity among HIV-1 variants poses serious challenges for vaccine development and for developing sensitive assays for screening, surveillance, diagnosis, and clinical management. Recognizing a need to develop a panel of HIV representing the current genetic and geographic diversity NIH/NIAID contracted the External Quality Assurance Program Oversight Laboratory (EQAPOL) to isolate, characterize and establish panels of HIV-1 strains representing global diverse subtypes and circulating recombinant forms (CRFs), and to make them available to the research community. HIV-positive plasma specimens and previously established isolates were collected through a variety of collaborations with a preference for samples from acutely/recently infected persons. Source specimens were cultured to high-titer/high-volume using well-characterized cryopreserved PBMCs from National y donors. Panel samples were stored as neat culture supernatant or diluted into defibrinated plasma. Characterization for the final expanded virus stocks included viral load, p24 antigen, infectivity (TCID), sterility, coreceptor usage, and near full-length genome sequencing. Viruses are made available to approved, interested laboratories using an online ordering application. The current EQAPOL Viral Diversity panel includes 100 viral specimens representing 6 subtypes (A, B, C, D, F, and G), 2 sub-subtypes (F1 and F2), 7 CRFs (01, 02, 04, 14, 22, 24, and 47), 19 URFs and 3 group O viruses from 22 countries. The EQAPOL Viral Diversity panel is an invaluable collection of well-characterized reagents that are available to the scientific community, including researchers, epidemiologists, and commercial manufacturers of diagnostics and pharmaceuticals to support HIV research, as well as diagnostic and vaccine development.Item Open Access Establishment and maintenance of a PBMC repository for functional cellular studies in support of clinical vaccine trials.(J Immunol Methods, 2014-07) Sambor, Anna; Garcia, Ambrosia; Berrong, Mark; Pickeral, Joy; Brown, Sara; Rountree, Wes; Sanchez, Ana; Pollara, Justin; Frahm, Nicole; Keinonen, Sarah; Kijak, Gustavo H; Roederer, Mario; Levine, Gail; D'Souza, M Patricia; Jaimes, Maria; Koup, Richard; Denny, Thomas; Cox, Josephine; Ferrari, GuidoA large repository of cryopreserved peripheral blood mononuclear cells (PBMCs) samples was created to provide laboratories testing the specimens from human immunodeficiency virus-1 (HIV-1) vaccine clinical trials the material for assay development, optimization, and validation. One hundred thirty-one PBMC samples were collected using leukapheresis procedure between 2007 and 2013 by the Comprehensive T cell Vaccine Immune Monitoring Consortium core repository. The donors included 83 human immunodeficiency virus-1 (HIV-1) seronegative and 32 HIV-1 seropositive subjects. The samples were extensively characterized for the ability of T cell subsets to respond to recall viral antigens including cytomegalovirus, Epstein-Barr virus, influenza virus, and HIV-1 using Interferon-gamma (IFN-γ) enzyme linked immunospot (ELISpot) and IFN-γ/interleukin 2 (IL-2) intracellular cytokine staining (ICS) assays. A subset of samples was evaluated over time to determine the integrity of the cryopreserved samples in relation to recovery, viability, and functionality. The principal results of our study demonstrate that viable and functional cells were consistently recovered from the cryopreserved samples. Therefore, we determined that this repository of large size cryopreserved cellular samples constitutes a unique resource for laboratories that are involved in optimization and validation of assays to evaluate T, B, and NK cellular functions in the context of clinical trials.Item Unknown Imaging system QA of a medical accelerator, Novalis Tx, for IGRT per TG 142: our 1 year experience.(Journal of applied clinical medical physics, 2012-07-05) Chang, Z; Bowsher, J; Cai, J; Yoo, S; Wang, Z; Adamson, J; Ren, L; Yin, FFAmerican Association of Physicists in Medicine (AAPM) task group (TG) 142 has recently published a report to update recommendations of the AAPM TG 40 report and add new recommendations concerning medical accelerators in the era of image-guided radiation therapy (IGRT). The recommendations of AAPM TG 142 on IGRT are timely. In our institute, we established a comprehensive imaging QA program on a medical accelerator based on AAPM TG 142 and implemented it successfully. In this paper, we share our one-year experience and performance evaluation of an OBI capable linear accelerator, Novalis Tx, per TG 142 guidelines.Item Unknown Introduction to a Special Issue of the Journal of Immunological Methods: Building global resource programs to support HIV/AIDS clinical trial studies.(J Immunol Methods, 2014-07) Sanchez, Ana M; Denny, Thomas N; O'Gorman, MauriceThis Special Issue of the Journal of Immunological Methods includes 16 manuscripts describing quality assurance activities related to virologic and immunologic monitoring of six global laboratory resource programs that support international HIV/AIDS clinical trial studies: Collaboration for AIDS Vaccine Discovery (CAVD); Center for HIV/AIDS Vaccine Immunology (CHAVI); External Quality Assurance Program Oversight Laboratory (EQAPOL); HIV Vaccine Trial Network (HVTN); International AIDS Vaccine Initiative (IAVI); and Immunology Quality Assessment (IQA). The reports from these programs address the many components required to develop comprehensive quality control activities and subsequent quality assurance programs for immune monitoring in global clinical trials including: all aspects of processing, storing, and quality assessment of PBMC preparations used ubiquitously in HIV clinical trials, the development and optimization of assays for CD8 HIV responses and HIV neutralization, a comprehensive global HIV virus repository, and reports on the development and execution of novel external proficiency testing programs for immunophenotyping, intracellular cytokine staining, ELISPOT and luminex based cytokine measurements. In addition, there are articles describing the implementation of Good Clinical Laboratory Practices (GCLP) in a large quality assurance laboratory, the development of statistical methods specific for external proficiency testing assessment, a discussion on the ability to set objective thresholds for measuring rare events by flow cytometry, and finally, a manuscript which addresses a framework for the structured reporting of T cell immune function based assays. It is anticipated that this series of manuscripts covering a wide range of quality assurance activities associated with the conduct of global clinical trials will provide a resource for individuals and programs involved in improving the harmonization, standardization, accuracy, and sensitivity of virologic and immunologic testing.Item Unknown Leukopak PBMC sample processing for preparing quality control material to support proficiency testing programs.(Journal of Immunological Methods, 2014-07) Garcia, Ambrosia; Keinonen, Sarah; Sanchez, Ana M; Ferrari, Guido; Denny, Thomas N; Moody, M AnthonyExternal proficiency testing programs designed to evaluate the performance of end-point laboratories involved in vaccine and therapeutic clinical trials form an important part of clinical trial quality assurance. Good clinical laboratory practice (GCLP) guidelines recommend both assay validation and proficiency testing for assays being used in clinical trials, and such testing is facilitated by the availability of large numbers of well-characterized test samples. These samples can be distributed to laboratories participating in these programs and allow monitoring of laboratory performance over time and among participating sites when results are obtained with samples derived from a large master set. The leukapheresis procedure provides an ideal way to collect samples from participants that can meet the required number of cells to support these activities. The collection and processing of leukapheresis samples require tight coordination between the clinical and laboratory teams to collect, process, and cryopreserve large number of samples within the established ideal time of ≤8 hours. Here, we describe our experience with a leukapheresis cryopreseration program that has been able to preserve the functionality of cellular subsets and that provides the sample numbers necessary to run an external proficiency testing program.Item Unknown Optimization and validation of a neutralizing antibody assay for HIV-1 in A3R5 cells.(Journal of immunological methods, 2014-07) Sarzotti-Kelsoe, Marcella; Daniell, Xiaoju; Todd, Christopher A; Bilska, Miroslawa; Martelli, Amanda; LaBranche, Celia; Perez, Lautaro G; Ochsenbauer, Christina; Kappes, John C; Rountree, Wes; Denny, Thomas N; Montefiori, David CA3R5 is a human CD4(+) lymphoblastoid cell line that was engineered to express CCR5 and is useful for the detection of weak neutralizing antibody responses against tier 2 strains of HIV-1. Here we describe the optimization and validation of the HIV-1 neutralizing antibody assay that utilizes A3R5 cells, performed in compliance with Good Clinical Laboratory Practice (GCLP) guidelines. The assay utilizes Renilla luciferase-expressing replication competent infectious molecular clones (IMC) encoding heterologous env genes from different HIV-1 clades. Key assay validation parameters tested included specificity, accuracy, precision, limit of detection and quantitation, specificity, linearity and range, and robustness. Plasma samples demonstrated higher non-specific activity than serum samples in the A3R5 assay. This assay can tolerate a wide range of virus input but is more sensitive to cell concentration. The higher sensitivity of the A3R5 assay in neutralization responses to tier 2 strains of HIV-1 makes it complementary to, but not a substitute for the TZM-bl assay. The validated A3R5 assay is employed as an endpoint immunogenicity test for vaccine-elicited neutralizing antibodies against tier 2 strains of HIV-1, and to identify correlates of protection in HIV-1 vaccine trials conducted globally.Item Unknown Optimization and validation of a neutralizing antibody assay for HIV-1 in A3R5 cells.(J Immunol Methods, 2014-07) Sarzotti-Kelsoe, M; Daniell, X; Todd, CA; Bilska, M; Martelli, A; LaBranche, C; Perez, LG; Ochsenbauer, C; Kappes, JC; Rountree, W; Denny, TN; Montefiori, DCA3R5 is a human CD4(+) lymphoblastoid cell line that was engineered to express CCR5 and is useful for the detection of weak neutralizing antibody responses against tier 2 strains of HIV-1. Here we describe the optimization and validation of the HIV-1 neutralizing antibody assay that utilizes A3R5 cells, performed in compliance with Good Clinical Laboratory Practice (GCLP) guidelines. The assay utilizes Renilla luciferase-expressing replication competent infectious molecular clones (IMC) encoding heterologous env genes from different HIV-1 clades. Key assay validation parameters tested included specificity, accuracy, precision, limit of detection and quantitation, specificity, linearity and range, and robustness. Plasma samples demonstrated higher non-specific activity than serum samples in the A3R5 assay. This assay can tolerate a wide range of virus input but is more sensitive to cell concentration. The higher sensitivity of the A3R5 assay in neutralization responses to tier 2 strains of HIV-1 makes it complementary to, but not a substitute for the TZM-bl assay. The validated A3R5 assay is employed as an endpoint immunogenicity test for vaccine-elicited neutralizing antibodies against tier 2 strains of HIV-1, and to identify correlates of protection in HIV-1 vaccine trials conducted globally.Item Unknown Optimized approach to decision fusion of heterogeneous data for breast cancer diagnosis.(Med Phys, 2006-08) Jesneck, Jonathan LeeAs more diagnostic testing options become available to physicians, it becomes more difficult to combine various types of medical information together in order to optimize the overall diagnosis. To improve diagnostic performance, here we introduce an approach to optimize a decision-fusion technique to combine heterogeneous information, such as from different modalities, feature categories, or institutions. For classifier comparison we used two performance metrics: The receiving operator characteristic (ROC) area under the curve [area under the ROC curve (AUC)] and the normalized partial area under the curve (pAUC). This study used four classifiers: Linear discriminant analysis (LDA), artificial neural network (ANN), and two variants of our decision-fusion technique, AUC-optimized (DF-A) and pAUC-optimized (DF-P) decision fusion. We applied each of these classifiers with 100-fold cross-validation to two heterogeneous breast cancer data sets: One of mass lesion features and a much more challenging one of microcalcification lesion features. For the calcification data set, DF-A outperformed the other classifiers in terms of AUC (p < 0.02) and achieved AUC=0.85 +/- 0.01. The DF-P surpassed the other classifiers in terms of pAUC (p < 0.01) and reached pAUC=0.38 +/- 0.02. For the mass data set, DF-A outperformed both the ANN and the LDA (p < 0.04) and achieved AUC=0.94 +/- 0.01. Although for this data set there were no statistically significant differences among the classifiers' pAUC values (pAUC=0.57 +/- 0.07 to 0.67 +/- 0.05, p > 0.10), the DF-P did significantly improve specificity versus the LDA at both 98% and 100% sensitivity (p < 0.04). In conclusion, decision fusion directly optimized clinically significant performance measures, such as AUC and pAUC, and sometimes outperformed two well-known machine-learning techniques when applied to two different breast cancer data sets.Item Unknown Protocol for fast scRNA-seq raw data processing using scKB and non-arbitrary quality control with COPILOT.(STAR protocols, 2022-12) Hsu, Che-Wei; Shahan, Rachel; Nolan, Trevor M; Benfey, Philip N; Ohler, UweWe describe a protocol to perform fast and non-arbitrary quality control of single-cell RNA sequencing (scRNA-seq) raw data using scKB and COPILOT. scKB is a wrapper script of kallisto and bustools for accelerated alignment and transcript count matrix generation, which runs significantly faster than the popular tool Cell Ranger. COPILOT then offers non-arbitrary background noise removal by comparing distributions of low-quality and high-quality cells. Together, this protocol streamlines the processing workflow and provides an easy entry for new scRNA-seq users. For complete details on the use and execution of this protocol, please refer to Shahan et al. (2022).Item Unknown Risk assessment and economic impact analysis of the implementation of new European legislation on radiopharmaceuticals in Italy: the case of the new monograph chapter Compounding of Radiopharmaceuticals (PHARMEUROPA, Vol. 23, No. 4, October 2011).(Curr Radiopharm, 2013-12) Chitto, Giuseppe; Di Domenico, Elvira; Gandolfo, Patrizia; Ria, Francesco; Tafuri, Chiara; Papa, SergioAn assessment of the new monograph chapter Compounding of Radiopharmaceuticals has been conducted on the basis of the first period of implementation of Italian legislation on Good Radiopharmaceuticals Practice (NBP) in the preparation of radiopharmaceuticals, in keeping with Decree by the Italian Ministry of Health dated March 30, 2005. This approach is well grounded in the several points of similarity between the two sets of regulations. The impact on patient risk, on staff risk, and on healthcare organization risk, has been assessed. At the same time, the actual costs of coming into compliance with regulations have been estimated. A change risk analysis has been performed through the identification of healthcare-associated risks, the analysis and measurement of the likelihood of occurrence and of the potential impact in terms of patient harm and staff harm, and the determination of the healthcare organization's controlling capability. In order to evaluate the economic impact, the expenses directly related to the implementation of the activities as per ministerial decree have been estimated after calculating the overall costs unrelated to NBP implementation. The resulting costs have then been averaged over the total number of patient services delivered. NBP implementation shows an extremely positive impact on risk management for both patients receiving Nuclear Medicine services and the healthcare organization. With regard to healthcare workers, instead, the implementation of these regulations has a negative effect on the risk for greater exposure and a positive effect on the defense against litigation. The economic impact analysis of NBP implementation shows a 34% increase in the costs for a single patient service. The implementation of the ministerial decree allows for greater detectability of and control over a number of critical elements, paving the way for risk management and minimization. We, therefore, believe that the proposed tool can provide basic criteria for analysis that could be used by other organizations setting about completing the same process.Item Unknown Setting objective thresholds for rare event detection in flow cytometry.(J Immunol Methods, 2014-07) Richards, Adam J; Staats, Janet; Enzor, Jennifer; McKinnon, Katherine; Frelinger, Jacob; Denny, Thomas N; Weinhold, Kent J; Chan, CliburnThe accurate identification of rare antigen-specific cytokine positive cells from peripheral blood mononuclear cells (PBMC) after antigenic stimulation in an intracellular staining (ICS) flow cytometry assay is challenging, as cytokine positive events may be fairly diffusely distributed and lack an obvious separation from the negative population. Traditionally, the approach by flow operators has been to manually set a positivity threshold to partition events into cytokine-positive and cytokine-negative. This approach suffers from subjectivity and inconsistency across different flow operators. The use of statistical clustering methods does not remove the need to find an objective threshold between between positive and negative events since consistent identification of rare event subsets is highly challenging for automated algorithms, especially when there is distributional overlap between the positive and negative events ("smear"). We present a new approach, based on the Fβ measure, that is similar to manual thresholding in providing a hard cutoff, but has the advantage of being determined objectively. The performance of this algorithm is compared with results obtained by expert visual gating. Several ICS data sets from the External Quality Assurance Program Oversight Laboratory (EQAPOL) proficiency program were used to make the comparisons. We first show that visually determined thresholds are difficult to reproduce and pose a problem when comparing results across operators or laboratories, as well as problems that occur with the use of commonly employed clustering algorithms. In contrast, a single parameterization for the Fβ method performs consistently across different centers, samples, and instruments because it optimizes the precision/recall tradeoff by using both negative and positive controls.Item Unknown Statistical methods for the assessment of EQAPOL proficiency testing: ELISpot, Luminex, and Flow Cytometry.(Journal of Immunological Methods, 2014-07) Rountree, Wes; Vandergrift, Nathan; Bainbridge, John; Sanchez, Ana M; Denny, Thomas NIn September 2011 Duke University was awarded a contract to develop the National Institutes of Health/National Institute of Allergy and Infectious Diseases (NIH/NIAID) External Quality Assurance Program Oversight Laboratory (EQAPOL). Through EQAPOL, proficiency testing programs are administered for Interferon-γ (IFN-γ) Enzyme-linked immunosorbent spot (ELISpot), Intracellular Cytokine Staining Flow Cytometry (ICS) and Luminex-based cytokine assays. One of the charges of the EQAPOL program was to apply statistical methods to determine overall site performance. We utilized various statistical methods for each program to find the most appropriate for assessing laboratory performance using the consensus average as the target value. Accuracy ranges were calculated based on Wald-type confidence intervals, exact Poisson confidence intervals, or via simulations. Given the nature of proficiency testing data, which has repeated measures within donor/sample made across several laboratories; the use of mixed effects models with alpha adjustments for multiple comparisons was also explored. Mixed effects models were found to be the most useful method to assess laboratory performance with respect to accuracy to the consensus. Model based approaches to the proficiency testing data in EQAPOL will continue to be utilized. Mixed effects models also provided a means of performing more complex analyses that would address secondary research questions regarding within and between laboratory variability as well as longitudinal analyses.Item Unknown Task Group 174 Report: Utilization of [18 F]Fluorodeoxyglucose Positron Emission Tomography ([18 F]FDG-PET) in Radiation Therapy.(Medical physics, 2019-10) Das, Shiva K; McGurk, Ross; Miften, Moyed; Mutic, Sasa; Bowsher, James; Bayouth, John; Erdi, Yusuf; Mawlawi, Osama; Boellaard, Ronald; Bowen, Stephen R; Xing, Lei; Bradley, Jeffrey; Schoder, Heiko; Yin, Fang-Fang; Sullivan, Daniel C; Kinahan, PaulThe use of positron emission tomography (PET) in radiation therapy (RT) is rapidly increasing in the areas of staging, segmentation, treatment planning, and response assessment. The most common radiotracer is 18 F-fluorodeoxyglucose ([18 F]FDG), a glucose analog with demonstrated efficacy in cancer diagnosis and staging. However, diagnosis and RT planning are different endeavors with unique requirements, and very little literature is available for guiding physicists and clinicians in the utilization of [18 F]FDG-PET in RT. The two goals of this report are to educate and provide recommendations. The report provides background and education on current PET imaging systems, PET tracers, intensity quantification, and current utilization in RT (staging, segmentation, image registration, treatment planning, and therapy response assessment). Recommendations are provided on acceptance testing, annual and monthly quality assurance, scanning protocols to ensure consistency between interpatient scans and intrapatient longitudinal scans, reporting of patient and scan parameters in literature, requirements for incorporation of [18 F]FDG-PET in treatment planning systems, and image registration. The recommendations provided here are minimum requirements and are not meant to cover all aspects of the use of [18 F]FDG-PET for RT.Item Unknown The Center for HIV/AIDS Vaccine Immunology (CHAVI) multi-site quality assurance program for cryopreserved human peripheral blood mononuclear cells.(J Immunol Methods, 2014-07) Sarzotti-Kelsoe, Marcella; Needham, Leila K; Rountree, Wes; Bainbridge, John; Gray, Clive M; Fiscus, Susan A; Ferrari, Guido; Stevens, Wendy S; Stager, Susan L; Binz, Whitney; Louzao, Raul; Long, Kristy O; Mokgotho, Pauline; Moodley, Niranjini; Mackay, Melanie; Kerkau, Melissa; McMillion, Takesha; Kirchherr, Jennifer; Soderberg, Kelly A; Haynes, Barton F; Denny, Thomas NThe Center for HIV/AIDS Vaccine Immunology (CHAVI) consortium was established to determine the host and virus factors associated with HIV transmission, infection and containment of virus replication, with the goal of advancing the development of an HIV protective vaccine. Studies to meet this goal required the use of cryopreserved Peripheral Blood Mononuclear Cell (PBMC) specimens, and therefore it was imperative that a quality assurance (QA) oversight program be developed to monitor PBMC samples obtained from study participants at multiple international sites. Nine site-affiliated laboratories in Africa and the USA collected and processed PBMCs, and cryopreserved PBMC were shipped to CHAVI repositories in Africa and the USA for long-term storage. A three-stage program was designed, based on Good Clinical Laboratory Practices (GCLP), to monitor PBMC integrity at each step of this process. The first stage evaluated the integrity of fresh PBMCs for initial viability, overall yield, and processing time at the site-affiliated laboratories (Stage 1); for the second stage, the repositories determined post-thaw viability and cell recovery of cryopreserved PBMC, received from the site-affiliated laboratories (Stage 2); the third stage assessed the long-term specimen storage at each repository (Stage 3). Overall, the CHAVI PBMC QA oversight program results highlight the relative importance of each of these stages to the ultimate goal of preserving specimen integrity from peripheral blood collection to long-term repository storage.Item Open Access The External Quality Assurance Oversight Laboratory (EQAPOL) proficiency program for IFN-gamma enzyme-linked immunospot (IFN-γ ELISpot) assay.(Journal of Immunological Methods, 2014-07) Sanchez, Ana M; Rountree, Wes; Berrong, Mark; Garcia, Ambrosia; Schuetz, Alexandra; Cox, Josephine; Frahm, Nicole; Manak, Mark; Sarzotti-Kelsoe, Marcella; D'Souza, M Patricia; Denny, Thomas; Ferrari, GuidoThe interferon-gamma enzyme-linked immunospot (IFN-γ ELISpot) assay has been developed and used as an end-point assay in clinical trials for infectious diseases and cancer to detect the magnitude of antigen-specific immune responses. The ability to compare data generated by different laboratories across organizations is pivotal to understand the relative potency of different therapeutic and vaccine strategies. We developed an external proficiency program for the IFN-γ ELISpot assay that evaluates laboratory performance based on five parameters: timeliness for data reporting; ability to handle cellular samples; detection of background (non-specific) responses; accuracy to consensus of the results; and precision of the measurements. Points are awarded for each criterion, and the sum of the points is used to determine a numeric and adjectival performance rating. Importantly, the evaluation of the accuracy to the consensus mean for the detection of antigen-specific responses using laboratory-specific procedures informs each laboratory and its sponsor on the degree of concordance of its results with those obtained by other laboratories. This study will ultimately provide the scientific community with information on how to organize and implement an external proficiency program to evaluate longitudinally the performance of the participating laboratories and, therefore, fulfill the requirements of the GCLP guidelines for laboratories performing end-point IFN-γ ELISpot assay for clinical trials.Item Open Access The Immunology Quality Assessment Proficiency Testing Program for CD3⁺4⁺ and CD3⁺8⁺ lymphocyte subsets: a ten year review via longitudinal mixed effects modeling.(Journal of Immunological Methods, 2014-07) Bainbridge, J; Wilkening, CL; Rountree, W; Louzao, R; Wong, J; Perza, N; Garcia, A; Denny, TNSince 1999, the National Institute of Allergy and Infectious Diseases Division of AIDS (NIAID DAIDS) has funded the Immunology Quality Assessment (IQA) Program with the goal of assessing proficiency in basic lymphocyte subset immunophenotyping for each North American laboratory supporting the NIAID DAIDS HIV clinical trial networks. Further, the purpose of this program is to facilitate an increase in the consistency of interlaboratory T-cell subset measurement (CD3(+)4(+)/CD3(+)8(+) percentages and absolute counts) and likewise, a decrease in intralaboratory variability. IQA T-cell subset measurement proficiency testing was performed over a ten-year period (January 2003-July 2012), and the results were analyzed via longitudinal analysis using mixed effects models. The goal of this analysis was to describe how a typical laboratory (a statistical modeling construct) participating in the IQA Program performed over time. Specifically, these models were utilized to examine trends in interlaboratory agreement, as well as successful passing of proficiency testing. Intralaboratory variability (i.e., precision) was determined by the repeated measures variance, while fixed and random effects were taken into account for changes in interlaboratory agreement (i.e., accuracy) over time. A flow cytometer (single-platform technology, SPT) or a flow cytometer/hematology analyzer (dual-platform technology, DPT) was also examined as a factor for accuracy and precision. The principal finding of this analysis was a significant (p<0.001) increase in accuracy of T-cell subset measurements over time, regardless of technology type (SPT or DPT). Greater precision was found in SPT measurements of all T-cell subset measurements (p<0.001), as well as greater accuracy of SPT on CD3(+)4(+)% and CD3(+)8(+)% assessments (p<0.05 and p<0.001, respectively). However, the interlaboratory random effects variance in DPT results indicates that for some cases DPT can have increased accuracy compared to SPT. Overall, these findings demonstrate that proficiency in and among IQA laboratories have, in general, improved over time and that platform type differences in performance do exist.Item Open Access VERITAS?: A time for VERIQAS™ and a new approach to training, education, and the quality assessment of CD4+ T lymphocyte counting (I).(Cytometry. Part B, Clinical cytometry, 2012-03) Barnett, David; Whitby, Liam; Wong, John; Louzao, Raul; Reilly, John T; Denny, Thomas NBackground
The aim of clinical laboratories is to produce accurate and reproducible results to enable effective and reliable clinical practice and patient management. The standard approach is to use both internal quality control (IQC) and external quality assessment (EQA). IQC serves, in many instances, as a "go, no go" tool to provide real time assurance that instruments and reagent or test systems are performing within defined specifications. EQA however, takes a snapshot at a specific point in time of the full testing process, results are compared to other laboratories performing similar testing but inevitably has some built in delay from sample issue to performance data review. In addition, if IQC or EQA identify areas of concern it can be difficult to determine the exact nature of the problem. In an attempt to address this problem, we have developed an instant QA panel that we have termed VERIQAS™, specifically for CD4(+) T lymphocyte counting, and have undertaken a "proof of principle" pilot study to examine how the use of VERIQAS™ could result in improvement of laboratory performance. In addition, we have examined how this approach could be used as a training and education tool (in a domestic/international setting) and potentially be of value in instrument validation/switch studies (a switch study being defined as a laboratory changing from one method/instrument to a new method/instrument with the VERIQAS™ panel being used as an adjunct to their standard switch study protocol).Methods
The basic panel consists of 20 stabilized samples, with predefined CD4(+) T lymphocyte counts, that span low clinically relevant to normal counts, including some blinded replicates (singlet up to quadruplicate combinations). The CD4(+) T lymphocyte target values for each specimen is defined as the trimmed mean ± 2 trimmed standard deviations, where the trimmed values are derived from the CD4(+) T lymphocyte counts reported by the participating centers (~780 laboratories) that receive each UK NEQAS for Leucocyte Immunophenotyping send out. Results for the VERIQAS™ panel were returned online, via a specially designed website, and the participant was provided with an immediate assessment (pass or fail).Results
To date, the panel has been preliminary trialed by eight laboratories to (i) assess pre-EQA qualification (two laboratories); (ii) address performance issues (two laboratories); or (iii) validate new instruments or techniques (four laboratories). Interestingly, even in this pilot study, the panel has been instrumental in identifying specific technical problems in laboratories with EQA performance issues as well as confirming that implementation of new techniques or instruments have been successful.Conclusion
We report here a new and novel "proof of principle" pilot study to quality assessment, that we have termed VERIQAS™, designed to provide instant feedback on performance. Participating laboratories receive 20 "blinded" samples that are in singlet up to quadruplicate combinations. Once a centre reports its results via a website, immediate feedback is provided to both the participant and the EQA organizers, enabling, if required, the initiation of targeted remedial action. We have also shown that this approach has the potential to be used as a tool for prequalification, troubleshooting, training and instrument verification. Pilot phase field trials with VERIQAS™ have shown that the panel can highlight laboratory performance problems, such as suboptimal instrument set up, pipetting and gating strategies, in a rapid and efficient manner. VERIQAS™ will now be introduced, where appropriate, as a second phase study within UK NEQAS for Leucocyte Immunophenotyping to assist those laboratories that have performance issues and also made available to laboratories for training and education of staff and instrument validation studies.