Performance Acceptance Criteria Research Articles

A-243 Quantitative Bacteriology Parameters for Seal-Packaged Agar Plates Challenged with Experimental Low-resource Tropical Field Clinic Stress

Abstract Background InTray® Mueller Hinton Chocolate (MHC) and Colorex™ Screen (CS) plates are in vitro diagnostic devices combining prepared bacteriological agar-media with Biomed’s ‘InTray®’ format (a double sealed-packaging cassette system that provides extended shelf-life 12 months past the date of manufacture). The devices are intended for use with clinical specimens to culture, isolate and identify potentially pathogenic bacteria. This study evaluated MHC and CS device performance at 7 and 13 months past the date of manufacture after product was experimentally challenged with stress replicating a low-resource tropical field clinic scenario. Methods MHC and CS plates were exposed to high temperature (40.5°C ± 0.5) and high humidity (74.5 ± 2.5% relative humidity) for an extended time-period (18 days). After the 18 days of heat treatment, the stress-group plates were stored in the lab at working room temperature (20-25°C) for the remainder of the study. Quantitative differences between control-groups (stored under refrigeration: 2-8°C) and heat-stress treatment groups for both MHC and CS were examined via CFU/mL measurements comparing three different lab-strain bacteria chosen based on clinical sub-culturing prevalence from blood bottles; Escherichia coli (ATCC 25922), Streptococcus pneumoniae (ATCC 6305), and Staphylococcus aureus (ATCC 25923). Briefly, 24-hour (18-30 h) old bacterial colony suspensions (grown on MHC) were prepared in 0.85% saline solution, adjusted to a 0.5 McFarland standard (here defined as 0.08-0.96 Absorbance @ 625 nm), measured in triplicate and diluted. A 20 μL volume from the lowest dilution (2.5 x 103 CFU/mL) was then spread-plated to each plate in order to count colony-forming units (CFU) in the acceptable range of 25-250. Results Quantitative CFU/mL differences between control-groups (stored under refrigeration: 2-8°C) and heat-stress-treatment groups (>40°C and high humidity) for MHC and CS for three different test bacteria (listed above) were not statistically significant when tested at 13 months. The CS stress and control group comparison for S. pneumoniae with a p value < 0.05 at 7 months had a p value > 0.05 at 13 months, suggesting that the difference between the CFU counts may not be significant in biological terms. Conclusions In conclusion, control and heat-stress treatment groups for both MHC and CS met acceptable performance criteria as expected for the products, 13 months after the date of manufacture.

The influence of different abiotic conditions on the concentrations of free and conjugated deoxynivalenol and zearalenone in stored wheat

Environmental factors influence fungal growth and mycotoxin production in stored grains. However, the concentrations of free mycotoxins and their conjugates and how they are impacted by different interacting environment conditions have not been previously examined. The objectives of this study were to examine the impact of storage conditions (0.93–0.98 aw) and temperature (20–25 °C) on (a) the concentrations of deoxynivalenol and zearalenone and their respective glucosides/conjugates and (b) the concentrations of emerging mycotoxins in both naturally contaminated and irradiated wheat grains inoculated with Fusarium graminearum. Contaminated samples were analysed for multiple mycotoxins using Liquid Chromatography Tandem Mass Spectrometry (LC–MS/MS). Method validation was performed according to the acceptable performance criteria set and updated by the European Commission regulations No. 2021/808/EC. As an important conjugate of deoxynivalenol, the concentrations of deoxynivalenol-3-glucoside were significantly different from its precursor deoxynivalenol at 0.93 aw (22% moisture content- MC) at 25 °C in the naturally contaminated wheat with a ratio proportion of 56:44% respectively. The high concentrations of deoxynivalenol-3-glucoside could be influenced by the wheat’s variety and/or harvested season/fungal strain type/location. Zeralenone-14-sulfate concentrations were surprisingly three times higher than Zearalenone in the naturally contaminated wheat at 0.98 aw (26% MC) at both temperatures. Emerging mycotoxins such as moniliformin increased with temperature rise with the highest concentrations at 0.95 aw and 25 °C. These findings highlight the influence and importance of storage aw x temperature conditions on the relative presence of free vs conjugated mycotoxins which can have implications for food safety.

Establishing Quality Assurance for HIV-1 Rapid Test for Recent Infection in Thailand through the Utilization of Dried Tube Specimens.

The present study focuses on establishing the quality assurance of laboratories for recent infections (RTRI) in Thailand. We developed a cold-chain independent method, using fully characterized plasma obtained from the Thai Red Cross Society, and prepared as dried tube specimens (DTS). Twenty microliters of HIV-seronegative, recent, and long-term infected samples were aliquoted into individual tubes and dried at room temperature, 20-30 degrees Celsius, in a biosafety cabinet overnight to ensure optimal preservation. The DTS external quality control and external quality assessment were tested for homogeneity and stability following the ISO/Guide 35 guidelines. The DTS panels were distributed to 48 sites (FY 2022) and 27 sites (FY 2023) across 14 and 9 provinces, respectively, in Thailand. The results from participating laboratories were collected and evaluated for performance. The results were scored, and acceptable performance criteria were defined as the proportion of panels correctly tested, which was set at 100%. The satisfactory performance ranged from 96% to 100% and was not significantly different among the 13 health regions. The developed and implemented DTS panels can be used to monitor the quality of RTRI testing in Thailand.

Systematic evaluation of the influence of Taq enzyme choice and amplification conditions on targeted environmental DNA assay performance and detection in field samples

AbstractTargeted environmental DNA (eDNA) studies mainly rely on quantitative real‐time polymerase chain reaction (qPCR) to amplify extremely low concentrations of DNA present in environmental samples. Understanding factors that influence targeted eDNA assay performance and detection in field samples, such as Taq DNA polymerase enzyme type and thermocycle protocol, is critical for the interpretation of qPCR results. We completed a systematic performance evaluation of five distinct targeted eDNA assays (eANFI6, eFISH1, eANBO5, eGLIN1, and eLICA3 targeting sablefish, general fish, Boreal toad, wood turtle, and the American bullfrog) by subjecting the same samples to analysis by five different Taq enzyme reagent mixes (Immolase, Environmental Master Mix (EMM), Amplitaq, QIAcuity, and QuantiNova) and the commonly used 2‐step (95°C, 60°C) and 3‐step (95°C, 64°C, 72°C) thermocycle protocols. We evaluated assay performance using a standardized dilution series of synthetic dsDNA target sequences and calculated limits of detection (LOD) and quantification (LOQ) and 95% confidence intervals for each combination of Taq enzyme and thermocycle protocol. All assays performed within acceptable performance criteria as defined by the Canadian national standard for targeted eDNA assays. Based on data generated by synthetic dsDNA fragments, the eDNA assays performed comparably regardless of the enzyme reagent mix and protocol used, except for eANFI6 and eGLIN1 using EMM and the 3‐step protocol, where no amplification was observed. On freshwater field samples, EMM and Immolase performed best. On marine field samples, Immolase, EMM, Qiacuity, and QuantiNova performed equally well, although EMM failed to amplify some samples. The work reveals that an enzyme reaction mix or thermocycle protocol can affect the result of an eDNA assay, but the appropriate choice also depends on the nature of the field sample. It is therefore imperative that these are considered when selecting appropriate reaction conditions and that they are clearly reported.

Thermal stability of carbon/polyimide coated optical fiber dried in hydrogen atmosphere

Manufacturing of polyimide-coated special optical fibers involves multiple steps of polyimide curing, including so called “drying” the fiber after draw. In this work, a fiber with carbon/polyimide coating is dried in a hydrogen atmosphere, and a complex analysis of the fiber characteristics (mechanical, optical, and thermal) is performed. Drying in hydrogen leads to an increase in strength of the carbon/polyimide coating and decreases a number of defects along the length of the fiber. The fiber strength analysis was performed in the framework of the Weibull distribution. No adverse effects of hydrogen on optical losses had been found at the operating wavelengths of the single-mode fiber at 1310 and 1550 nm. The temperature stability has been analyzed via thermogravimetry for the fiber samples taken before and after the drying in air and hydrogen environments. The activation energies were calculated by Kissinger method. Per the performance acceptance criteria, the upper use temperature of the hydrogen atmosphere dried coating reach 397 °C, while for the air-dried fiber it is just 215 °C. Thus, drying carbon/polyimide coated optical fibers in hydrogen atmosphere provides a promising improvement, which allows further use of these fibers in harsh environments.

B-304 Performance of Alternative Proficiency Testing Program for Bayesian Forecasting Tool of Infliximab Exposure in Clinical Pharmacokinetic Laboratory

Abstract Background We have developed a Bayesian (PK) forecasting and dosing tool for Infliximab (IFX), a monoclonal antibody targeting Tumor necrosis factor α. This novel dosing tool employs machine learning and proficiency of the algorithm must be established. Our objective was to implement an alternative proficiency testing program (APT) in the routine clinical (PK) laboratory. Methods The test (PredictrPK® IFX) is validated and measures serum IFX and antibodies to IFX (ATI) concentrations using a homogenous mobility shift assay by size exclusion HPLC and albumin (ALB) by nephelometry. Determinations are inputted with weight, dose, and interdose interval provided on test requisition in the Bayesian forecasting PK tool which uses non-linear mixed effect to calculate the individual parameter estimates, Clearance and predicted trough concentrations. APT is conducted using five previously tested patient samples, blinded to testing personnel and retested in the same manner as the routine method of analysis every six months. APT consists of two components, a component evaluating standalone IFX, ATI, and ALB analytes, and a second component evaluating the Bayesian forecasting PK tool in estimating Clearance and trough concentrations. Criteria for acceptable performance is set at target value ± 20% for standalone analytes and forecasted PK estimates. Acceptable ATP score corresponds to 4/5 samples with passing grade. Results The results from two consecutive APT results with six months intervals is presented in Fig. 1. Comparing the results to the original assayed data at the time of collection, the APT results showed IFX concentration, ATI and ALB within 20% of target value. Clearance and estimated trough concentrations calculated using the Bayesian forecasting tool also met acceptability criteria (100% passing grade). Conclusion We have implemented an APT for Bayesian forecasting tool in the clinical PK laboratory.

Development and validation of a liquid chromatographic-tandem mass spectrometric assay for the quantification of the direct acting antivirals glecaprevir and pibrentasvir in plasma

BackgroundDirect acting antiviral (DAA) therapies are effective in the treatment and management of chronic HCV infections. Glecaprevir (GLE) and pibrentasvir (PIB) are pangenotypic DAAs that are delivered alone or as a fixed-dose oral formulation to treat chronic HCV infections with or without cirrhosis. Sensitive and dynamic bioanalytical assays are needed to understand the pharmacology of GLE and PIB. MethodsDrug free K2EDTA plasma was spiked with GLE, PIB, and their internal standards. Drugs were extracted from plasma via protein precipitation, and subsequently quantified using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The method was validated according to regulatory recommendations, and evaluated in remnant plasma samples from individuals prescribed GLE and PIB. ResultsThe analytical measuring ranges for GLE and PIB were 0.25–2000 ng/mL and 0.25–1000 ng/mL, respectively. The method showed acceptable accuracy and precision for both DAAs. GLE and PIB in plasma were stable following six freeze thaw cycles and at room temperature for up to 67 h. All analytes were stable in whole blood incubated at room temperature for 24 h, and at 40 °C and 100% humidity for 2 h. Negligible percent matrix effects were observed for PIB and PIB-IS across the measuring range of the assay. Significant ion suppression was observed for GLE, with an average matrix effects of 27.9%. However, relative matrix effects were < 6.3% between drug and internal standard, and deemed acceptable. Assay validation assessments in alternative matrices also met acceptance criteria. Both DAAs were successfully measured in remnant plasma samples from individuals administered GLE and PIB. ConclusionsAn LC-MS/MS method for GLE and PIB quantification in plasma has been developed and validated. The assay met acceptable performance criteria and may be used in downstream applications to characterize DAA pharmacology for HCV treatment.

Screening for fetal alcohol spectrum disorder in infants and young children.

Introduction: With an estimated prevalence of up to five percent in the general population, fetal alcohol spectrum disorders (FASD) are the most common neurodevelopmental disorder and more prevalent than autism. Early identification and subsequent early intervention have the potential to improve developmental trajectory of children with FASD. In addition, new research suggests supplementation with choline may ameliorate the developmental impairments associated with prenatal alcohol exposure. Availability of a screening tool with acceptable epidemiologic performance criteria may be clinical useful in identification of young children at increased risk for FASD. In this paper we describe the Early Fetal Alcohol Spectrum Disorder Screening Test (E-FAST) to identify young children at increased risk for an FASD. Methods: We developed the E-FAST dataset from previously published studies, comprised of 281 children under 5years of age, 180 (64.1%) were diagnosed with FASD and 101 (35.9%) were non-FASD. Analysis: The analysis identified seven useful variables (prenatal alcohol exposure, ADHD (Attention Deficit Hyperactivity Disorder), foster care or adopted, small OFC (occipital frontal circumference), communication impairments, impaired social skills, and cognitive deficits. All variables were categorized as yes/no for ease of use in a screening tool. Risk ratios for each of the seven indicators were estimated using two-way table analyses. Weights for each variable were estimated based on the relative strength of their odds ratios. Results: The average age was 2.7years of age (S.D. 1.29) and ranged from infant (6.4%) to 4years old (35.9%). Maternal alcohol use alone had a sensitivity of 0.97, specificity 0.65, and accuracy 0.86. For the combined seven variables, sensitivity was 0.94, specificity 0.74, and accuracy 0.87. Thus, the seven-item E-FAST screen had acceptable epidemiologic screening characteristics. Discussion: In the United States, up to 547 infants with FASD are born each day which far exceeds the capacity of multidisciplinary diagnostic clinics. During routine clinical management of infants and young children the use of an evidence-based screening tool provides a time efficient means to exclude large numbers of young children from further follow-up for FASD. Conversely, a positive screen identifies a smaller number of children at increased risk for FASD requiring more intensive evaluation and follow-up.

#5228 ASSESSING ACCURACY OF POINT-OF-CARE CREATININE AND POTASSIUM TESTING IN A MULTI-ETHNIC SOUTH LONDON POPULATION

Abstract Background and Aims Point-of-care (POC) testing allows rapid analysis of blood samples without delay of laboratory testing. For patients with chronic kidney disease (CKD), this allows for instant identification of changes in kidney function, such as acute kidney injury or hyperkalaemia, and assists in timely decision-making in outpatient or community settings (e.g. commencing or up-titrating medications dependent on renal function). It also provides an opportunity for early recognition of CKD in high-risk populations by providing screening in more convenient and trusted settings. To achieve these aims, it essential to confirm that POC systems can produce accurate results in real world settings with diverse patient populations. This pilot validation study assessed accuracy of the Siemens Epoc Blood Analysis System venous creatinine and potassium compared with laboratory assays in a diverse UK renal patient population. Method Venous blood samples from 57 patients, aged ≥ 18 years old and receiving care at a London teaching hospital, were analysed using the Siemens Epoc Blood Analysis System. POC-Creatinine (POC-Cr) and Potassium (POC-K) measurements were compared with laboratory (IDMS-traceable Siemens enzymatic and Roche) assays. Demographics (age, ethnicity and sex) were recorded for each patient. Passing-Bablock regression analysis were used to compare the results using both methods. Comparison between testing methods was assessed using a Bland-Altman plot. Limits of agreement were compared with CLIA criteria for acceptable performance for creatinine and potassium. Results The mean age was 55.6 ± 15.3 years old and 64.9% were male (n = 37). The majority of patients were of White (N = 23; 40.4%), Black (N = 21; 36.8%) or South Asian (N = 9; 15.8%) ethnicity respectively. Median POC-Cr was 216 μmol/L (interquartile range (IQR) 111, 396 μmol/L) versus median venous creatinine 212 μmol/L (IQR 114, 380 μmol/L). The median bias for creatinine was +1.0 (IQR -3.0, +8.0). Limits of agreement on Bland-Altman plot were +36.5μmol/L and – 34.4μmol/L (Figure 1). Overall, there remained a strong positive correlation between POC-Cr and IDMS-traceable enzymatic creatinine assays (R = 0.997; P&amp;lt;0.0001) (Figure 2). Median POC-K was 4.3 mmol/L, IQR 4.0, 4.8 mmol/L versus median venous potassium 5.0, IQR 4.3, 5.3 mmol/L. The median bias for potassium was +0.4 (IQR +0.2, +0.5). Limits of agreement on Bland-Altman plot were +0.16mmol/L and – 0.91mmol/L (Figure 3). A strong positive correlation between POC-K and laboratory potassium assays was also confimed (R = 0.937; P&amp;lt;0.0001) (Figure 4). Conclusion There was strong positive correlation between POC and laboratory analysis of venous creatinine, comparable to product literature (R = 0.997 vs 0.998). Limits of agreement were outside the CLIA criteria for acceptable performance (+/- 27μmol/L). POC and laboratory potassium levels were also strongly correlated and comparable to product literature (R = 0.937 vs 0.997). Limits of agreement were outside the CLIA criteria for acceptable performance (+/- 0.5mmol/L). This discrepancy is likely due to haemolysis in some blood samples and small sample size. Overall, this pilot study demonstrated a strong correlation between POC and laboratory testing. Larger scale evaluation is still required in a multi-ethnic population, prior to assessing feasibility in community-led protocolized pathways.

Evaluation of Measurement Performance of Routine Chemistry Analytes by Application of Sigma Metrics, Method Evaluation Decision (MEDx) Charts and Quality Goal Index (QGI)

Objective: To evaluate performance of routine chemistry analytes in a tertiary care hospital laboratory by the application of sigma metrics. Introduction: Six sigma (6σ) is a popular Quality Management System (QMS) tool. Laboratories are increasingly using the six sigma method for the objective assessment and comparison of the analytical methods and instrument performance. Six sigma is about measuring or counting the number of defects. It quantifies the performance of a process as a rate of Defects-Per-Million-Opportunities (DPMO or DPM). The aim is to assess the performance and to eliminate or reduce the variation in a process. Materials and Methods: This prospective study was conducted over a period of six months duration. Sigma metrics were calculated using coefficient of variation (%CV), %Bias and total allowable error (%TEa). For %CV we used Internal Quality Control (IQC) samples, at two levels L1 and L2. Daily IQC results of L1 and L2 for 25 routine chemistry analytes were recorded in an excel sheet and %CV was calculated for each analyte for the period of study. For each analyte %Bias was calculated based on values obtained from monthly RIQAS-EQA program data. The total allowable error (%TEa) values for each analyte were extracted from various sources like Clinical Laboratories Improvement Amendment act (CLIA), Canadian Fixed Limits from the College of Physicians and Surgeons of Saskatchewan (CFX) and Spanish Society of Clinical Chemistry and Molecular Pathology (SEQC) table of Desirable Quality Specifications based on Biological Variation (BV) criteria for acceptable performance. Sigma values were calculated. The minimal acceptable performance criteria was considered as 3 sigma. Normalized MEDx charts were used to plot sigma metrics to visually present the performance. Quality Goal Index (QGI) analysis was carried out as a part of Root Cause Analysis (RCA). Results: Highest sigma value of 16.7 was noted for HDL-C and the lowest of 2.08 for chloride at level L1. Many analytes like ALP, Amylase, AST, CK, GGT, HDL-C, Magnesium and Uric Acid attained world class quality performance at both levels L1 and L2 with sigma levels of &gt;6. Many other analytes showed satisfactory performance with sigma levels of &gt;3. Sodium, potassium, chloride and urea did not show a satisfactory performance. Conclusion: Sigma metrics evaluation of analytical performance of our laboratory showed an acceptable performance for a wide range of analytes in patient samples.

IPVS policy statement on HPV nucleic acid testing guidance for those utilising/considering HPV as primary precancer screening: Quality assurance and quality control issues.

We advise that only clinically validated HPV assays which have fulfilled internationally accepted performance criteria be used for primary cervical screening. Further, assays should be demonstrated to be fit for purpose in the laboratory in which they will ultimately be performed, and quality materials manuals and frameworks will be helpful in this endeavor. Importantly, there is a fundamental shortage of well validated, low-cost, low complexity HPV tests that have demonstrated utility in a near-patient setting; representing a significant challenge and focus for future development in order to reach the WHO's goal of eliminating cervical cancer.

Six Sigma metrics as an indicator for Randox International Quality Assessment Scheme in clinical biochemistry laboratory

Context: Six Sigma methodology represents an evolution in quality assessment and management that has been implemented widely in industry and health sector since the mid-1980s. It consists of five steps: define, measure, analyze, improve, and control (DMAIC). Randox International Quality Assessment Scheme (RIQAS) program Target Score (TS) is a performance indicator, and Sigma metrics is considered as a gold standard for overall performance. Aim: The aim of this study is to present the TS and Sigma metrics observed in clinical chemistry laboratory in SSK Hospital associated with Lady Hardinge Medical College to evaluate the analytical performance of T3, T4, and thyroid-stimulating hormone (TSH) parameters. The study period is from June to August 2022. Materials and Methods: The Sigma metrics for the various analytes was calculated by the following equation: Σ = TEa − bias/CV (TEa—total allowable error, CV—coefficient of variation). Acceptable performance criteria: standard deviation index (SDI) &lt;2; TS &gt;50; the closer a value is to 120, the better the performance, &lt;40 is unacceptable; and %deviation should be less than the defined acceptable limits which is unique to each analyte, the closer the value is to zero, the better is the performance. Results: Our RIQAS results lie in acceptable range except for T3 in the month of August in which TS was unacceptable. Our laboratory Sigma metrics were below three sigmas. The findings of our exercise emphasize the need for a detailed evaluation, and we promote the use of Sigma metrics in quality assurance in the best interest of the patient. Conclusion: Quality check strategies were applied according to the TS and sigma values.

Development and validation of a primary IDMS method for the quantification of 12 sulfonamides in meat by using LC-MS/MS

A new liquid chromatography-tandem mass spectrometry (LC-IDMS) method was developed and validated for a simultaneous multi residue analysis of 12 sulfonamides (SAs) in beef meat. Sulfonamides were isolated from meat with a solvent extraction procedure. Reliable quantitative evaluation was accomplished by using each of the sulfonamides as the internal standart of their own isotopes. Matrix-matched calibration curves were used. Studied performance characteristics were linearity, recovery, precision, detection and quantification limits and robustness. Results were compatible to method performance acceptance criteria according to UME. Recovery results were 82-116%. Measurement uncertainty was calculated from “top down” approach”. Relative measurement uncertainty was between 7-14%.

Improving the quality of HIV rapid testing in Ghana using the dried tube specimen-based proficiency testing program.

BackgroundThe introduction of human immunodeficiency virus (HIV) antibody rapid testing (RT) in resource-limited settings has proven to be a successful intervention to increase access to prevention measures and improve timely linkage to care. However, the quality of testing has not always kept pace with the scale-up of this testing strategy. To monitor the accuracy of HIV RT test results, a national proficiency testing (PT) program was rolled out at selected testing sites in Ghana using the dried tube specimen (DTS) approach.MethodsBetween 2015 and 2018, 635 HIV testing sites, located in five regions and supported by the U.S. President’s Emergency Plan for AIDS Relief (PEPFAR), were enrolled in the HIV PT program of the Ghana Health Service National AIDS/STI Control Programme. These sites offered various services: HIV Testing and Counselling (HTC), prevention of mother-to-child transmission (PMTCT) and Antiretroviral Treatment (ART). The PT panels, composed of six DTS, were prepared by two regional laboratories, using fully characterized plasma obtained from the regional blood banks and distributed to the testing sites. The results were scored by the PT providers according to the predefined acceptable performance criteria which was set at ≥ 95%.ResultsSeven rounds of PT panels were completed successfully over three years. The number of sites enrolled increased from 205 in round 1 (June 2015) to 635 in round 7 (December 2018), with a noticeable increase in Greater Accra and Eastern regions. The average participation rates of enrolled sites ranged from 88.0% to 98.0% across the PT rounds. By round 7, HTC (257/635 (40.5%)) and PMTCT (237/635 (37.3%)) had a larger number of sites that participated in the PT program than laboratory (106/635 (16.7%)) and ART (12/635 (1.9%)) sites. The average testing performance rate improved significantly from 27% in round 1 to 80% in round 7 (p < 0.001). The highest performance rate was observed for ART (100%), HTC (92%), ANC/PMTCT (90%) and Laboratory (89%) in round 5.ConclusionThe DTS PT program showed a significant increase in the participation and performance rates during this period. Sub-optimal performances observed was attributed to non-compliance to the national testing algorithm and testing technique. However, the implementation of review meetings, peer-initiated corrective action, supportive supervisory training, and mentorship proved impactful. The decentralized approach to preparing the PT panels ensured ownership by the region and districts.

Using high-sensitivity cardiac troponin for the exclusion of inducible myocardial ischemia in patients without previously known coronary artery disease

Abstract Background The rapid and safe exclusion of functionally relevant coronary artery disease (CAD) is a crucial, yet unmet clinical need. High-sensitivity cardiac troponin (hs-cTn) may be an attractive strategy, particularly in patients without previously known CAD. Purpose To derive and internally validate optimal rule-out cutoffs for an early and safe exclusion of functionally relevant CAD in symptomatic patients without previously known CAD. Methods In an ongoing single-center, prospective, cohort study, we enrolled consecutive patients without previously known CAD that were referred with symptoms possibly related to functionally relevant CAD. Cardiac troponin concentrations were measured at presentation using two high-sensitivity assays (Elecsys hs-cTnT and Architect hs-cTnI). Presence of functionally relevant CAD was adjudicated by 2 independent cardiologists, blinded to hs-cTn measurements, using MPI-SPECT/CT in all patients, as well as coronary angiography and fractional flow reserve measurements, whenever available. The primary diagnostic outcome was safety for early rule-out of functionally relevant CAD, quantified by sensitivity and the negative predictive value (NPV). The co-primary prognostic outcomes were cumulative incidences of cardiovascular death and all-cause death after 5 years. A NPV ≥90% and sensitivity ≥90% were predefined as acceptable performance criteria. The derived cutoffs were further evaluated in pre-specified subgroups. Internal validity was assessed with a bootstrapping procedure for a realistic estimate in similar future patients. Cumulative incidence curves stratified by the presence of functionally relevant CAD and hs-cTn concentrations below and above the derived cutoffs were constructed. Results Among 2111 eligible patients, 498 (23.6%) had a final diagnosis of functionally relevant CAD. Median age was 68 years and 938 (44.4%) were female. For ruling out functionally relevant CAD, a hs-cTnT concentration &amp;lt;5 ng/L resulted in a sensitivity of 90.8% (95% CI: 87.9–93.0%) and a NPV of 90.2% (95% CI: 87.1–92.5), triaging 468 (22.2%) patients towards rule-out. Similarly, a hs-cTnI concentration &amp;lt;2 ng/L resulted in a sensitivity of 91.6% (95% CI: 88.8–93.7%) and a NPV of 90.0% (95% CI: 86.8–92.6), triaging 422 (20.0%) patients. Internal validation showed robustness of these findings. The diagnostic performance of the derived cutoffs did not significantly vary across the subgroups. Hs-cTn concentrations above the derived cutoffs were associated with a substantially higher cumulative event rate of cardiovascular death (hs-cTnT: 7.0% vs. 0.8%; hs-cTnI: 6.6% vs. 1.2%) and all-cause death (hs-cTnT: 14.3% vs. 2.4%; hs-cTnI: 13.1% vs. 4.4%) during 5-years follow-up (log rank p&amp;lt;0.001 for all). Conclusion In symptomatic patients without previously known CAD, very low hs-cTn concentrations may generally allow to safely and effectively exclude functionally relevant CAD. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Swiss National Science FoundationSwiss Heart Foundation

Innovative Techniques for Optimization of Far-Field Diverter Materials to Enhance Fracture Geometry in Carbonate Reservoirs

Summary The technique of employing specialized particulates for far-field diversion is well established during hydraulic fracturing treatments in unconventional formations and is being investigated for use in conventional formations. Far-field diverters (FFD) divert fluid away from the wellbore far into the formation. The injection of FFD at the beginning of the treatment provides an additional stress barrier between the producing interval and adjacent layers by depositing at the layer boundaries where a higher leakoff is encountered. The ensuing restriction in height growth maximizes fracture extension within the producing zone, optimizing geometry for increased hydrocarbon production while limiting excess water. Polylactic acid (PLA) polymer is self-degradable, compatible with reservoir fluids, and has a variety of compositions for different temperature applications. Blending proppant with PLA has been seen to significantly improve the strength of the deposited FFD. Therefore, PLA powder and silica proppant are blended to develop Generation-1 FFD (FFD-Gen1). However, many silica proppants have greater density than PLA, leading to separation during transport which prevents these two components from depositing evenly at the upper fracture boundary. This results in a situation in which excessive downward growth is prevented while upward growth is left unchecked. For this reason, both components need to be simultaneously deposited to develop an effective seal. Generation-2 FFD (FFD-Gen2) is developed by replacing the silica proppant of FFD-Gen1 with a deformable proppant having a density nearly equal to the polymer, which enables uniform deposition on all adjacent formation boundaries where leakoff is encountered. The deformable characteristic improves the pressure withstanding capacity of the diverter pack. The deposition and degradation behaviors are investigated in the laboratory by performing high-temperature high-pressure (HTHP) filter press and plug stability experiments. Experimental findings suggest that the primary selection criteria for acceptable performance are the material’s mechanical properties. This methodology is used to select the appropriate FFD materials to optimize fracture geometry in carbonate reservoirs. Successful applications prevent excessive water production and substantially increase hydrocarbon production as illustrated in a three-well case study.

Serum Irisin: A Potential Diagnostic Marker for Insulin Resistance in Acne Vulgaris.

Acne vulgaris (AV) is a chronic inflammatory disease of the pilosebaceous unit. Many factors are involved in the occurrence of acne. It has been confirmed that some adipokines play an important role in the development of AV. Irisin is a novel adipokine, which is highly expressed in skeletal muscle, liver, and fat. It improves insulin resistance (IR) by inducing the browning of white adipose tissue, increasing heat production and energy expenditure. The purpose of this study was to investigate the role of serum irisin as an adipokine to explore its function in the pathogenesis of AV and its correlation with IR, and whether it can be used as a potential biomarker of insulin sensitivity. Although the hyperinsulinemic-euglycemic clamp remains the gold standard for accurate determination of IR, it cannot be performed routinely. Various alternative simpler measures have been used, the most common being homeostasis model assessment. However, these metrics are limited by their accuracy, cost, and blood collection requirements.[1] Therefore, an effective and feasible serum biomarker is an attractive and relatively straightforward method, which may provide clinicians with a more accurate and simple method for the prediction and diagnosis of IR. IR can often be detected before other symptoms appear, so establishing an early diagnosis method will allow for the appropriate treatment of patients before the disease develops. The study included 171 subjects; 115 patients with newly diagnosed AV and 56 apparently healthy subjects. The contents of irisin and interleukin-1 alpha in serum were determined by enzyme-linked immunosorbent assay. The IR index was calculated by the homeostasis model. Serum irisin levels in AV patients and control group were (24.0 ± 11.3) and (104.3 ± 27.0) ng/dl, respectively, which were significantly lower than those in control group (P < 0.001). Serum irisin was negatively correlated with IR (r = -0.711, P 0.001). The sensitivity of irisin was 100.0%, the specificity was 92.8%, and the cutoff point was 53.32. The decrease of serum irisin level could predict the patients with IR in acne. Serum irisin levels in AV patients were significantly decreased. Serum irisin showed acceptable performance criteria in the diagnosis of AV with IR. Serum irisin seems to be a good diagnostic and prognostic marker for IR. Further multi-center studies are needed to confirm this link, which could pave the way for new treatment options.

Atellica CH 930 chemistry analyzer versus Cobas 6000 c501 and Architect ci4100 - a multi-analyte method comparison

Abstract Large clinical laboratories often rely on multiple chemistry analyzers. However, when a new analyzer is introduced, the laboratory must establish whether the old and new methods are comparable and can be used interchangeably. In this study, we compared the newly introduced Atellica CH930 chemistry analyzer with the already established Architect ci4100 and Cobas 6000 c501 from our laboratory. Patient samples were randomly selected from daily routine testing and a total of 22 analytes were investigated. Total error (TEobs) between test (Atellica) and comparative (Architect and Cobas) methods was calculated at relevant medical decision levels (MDL). For demonstrative purposes, the assessment of method comparability was based on three different criteria: allowable total error (TEa) derived from biological variation (BV), CLIA proficiency testing criteria for acceptable analytical performance, and CLIA-calculated Sigma metrics. These sets of analytical performance specifications were also compared, and their strengths and limitations are discussed in this paper. Performance of Atellica CH930 against Architect ci4100 was acceptable or nearly acceptable at 82%, 95%, and 64% of the 22 investigated MDLs across 9 analytes, according to BV-TEa, CLIA-TEa, and CLIA-calculated Sigma metrics, respectively. Similarly, performance of Atellica CH930 against Cobas 6000 c501 was acceptable or nearly acceptable at 61%, 93%, and 63% of the 54 investigated MDLs across 22 analytes, according to BV-TEa, CLIATEa, and CLIA-calculated Sigma metrics, respectively. However, method comparability should not be evaluated by a “one size fits all” approach as some analytes require different criteria of acceptability, ideally based on medically allowable error and clinical outcome.

Rapid Identification and Antimicrobial Susceptibility of Gram Negative Bacteria Detected in Positive Blood Culture Bottles

Abstract Background Blood stream infection (BSI) is one of the most serious situations in infectious disease .Accurate and timely identification of the causative agent and determination of its antimicrobial susceptibility profile are essential for guiding targeted and effective antimicrobial treatment. Current methods involve culturing of blood in a liquid medium and subsequently subculturing of signal positive bottles on solid media in order to obtain isolated colonies that can be further used in identification and susceptibility testing of the isolates. The standard method requires additional 48-72hours following the appearance of a positive signal in order to provide a reliable patient report. On the other hand, rapid identification of the causative agent and determination of its susceptibility profile by direct inoculation of the biochemical test media with the blood-broth mixtures from signal positive bottles and performing primary susceptibility testing might help reducing the time needed for provision of results compared to the standard isolated-colony based method and hence would help the rapid initiation of effective and targeted antimicrobial therapy and reduce the bacteremia-related morbidity and mortality. Objective The aim of the present study was to determine the accuracy and precision of the non-standard methods (direct identification and susceptibility testing using the blood/broth mixture) by comparing its results to those of the standard isolate-based identification and susceptibility testing methods. Material and method The study included 52 signal blood culture bottles yielding gram negative isolates. Bottles were selected amongst blood culture bottles submitted to the Main Microbiology laboratory, Ain Shams University hospital, for culture and antimicrobial susceptibility testing during the period between May 2018 and October 2018. a portion of the blood-broth mixture was aspirated from positive blood culture bottles, after being well mixed, and was subcultured onto agar media for the isolation of the causative agent and subsequently determination of its antimicrobial susceptibility profile was performed. Another part of the aspirated blood-broth mixture was diluted with sterile saline, its turbidity was adjusted against a 0.5McFarland standard and was used to inoculate directly the biochemical test media panel used for the identification of gram negative organisms as well as to perform direct (primary) antimicrobial susceptibility. Results The present study revealed there was 100% categorical agreement between the results of the direct biochemical inoculation method and those of the standard isolate-based inoculation method regarding the identification of the causative agent. The results of the direct biochemical identification method were also consistent giving rise to a 100% withinrun precision categorical agreement and a 100% between-run precision categorical agreement. The overall categorical agreement between the results of the standard isolate-based AST method and the results of the direct (primary susceptibility) AST method was 96.3% for the signal blood culture media. The major error rate was 0.5% whereas the minor error rate was 3.3% . Consistent results were also obtained for the AST done directly from the signal blood culture bottles since the between-run and within-run precision categorical agreement were 96.3% and 98.6%, respectively. Conclusion the overall performance of the AST done directly from positive blood culture bottles fulfilled the acceptable performance criteria specified in the Cumitech 31A so the direct method can be used for the earlier determination of AST and identification of Gram negative bacteria and thus to reduce the time for early initiation of appropriate antibiotic

Physiologically-Based Pharmacokinetic Modelling of Entrectinib Parent and Active Metabolite to Support Regulatory Decision-Making.

Entrectinib is a selective inhibitor of ROS1/TRK/ALK kinases, recently approved for oncology indications. Entrectinib is predominantly cleared by cytochrome P450 (CYP)3A4, and modulation of CYP3A enzyme activity profoundly alters the pharmacokinetics of both entrectinib and its active metabolite M5. We describe development of a combined physiologicallybased pharmacokinetic (PBPK) model for entrectinib and M5 to support dosing recommendations when entrectinib is co-administered with CYP3A4 inhibitors or inducers. A PBPK model was established in Simcyp® Simulator. The initial model based on in vitro-in vivo extrapolation was refined using sensitivity analysis and non-linear mixed effects modeling to optimize parameter estimates and to improve model fit to data from a clinical drug-drug interaction study with the strong CYP3A4 inhibitor, itraconazole. The model was subsequently qualified against clinical data, and the final qualified model used to simulate the effects of moderate to strong CYP3A4 inhibitors and inducers on entrectinib and M5 pharmacokinetics. The final model showed good predictive performance for entrectinib and M5, meeting commonly used predictive performance acceptance criteria in each case. The model predicted that co-administration of various moderate CYP3A4 inhibitors (verapamil, erythromycin, clarithromycin, fluconazole, and diltiazem) would result in an average increase in entrectinib exposure between 2.2- and 3.1-fold, with corresponding average increases for M5 of approximately 2-fold. Co-administration of moderate CYP3A4 inducers (efavirenz, carbamazepine, phenytoin) was predicted to result in an average decrease in entrectinib exposure between 45 and 79%, with corresponding average decreases for M5 of approximately 50%. The model simulations were used to derive dosing recommendations for co-administering entrectinib with CYP3A4 inhibitors or inducers. PBPK modeling has been used in lieu of clinical studies to enable regulatory decision-making.