Passing-Bablok Regression Analysis Research Articles

Comparing Dried Blood Spots and Plasma Concentrations for Busulfan Therapeutic Drug Monitoring in Children.

Busulfan (Bu) is one of the conditioning regimen components for pediatric hematopoietic stem cell transplantation. Bu therapeutic drug monitoring (TDM) is essential for a successful treatment outcome and toxicity evasion. Dried blood spot (DBS) sampling is a rapid and simple method for Bu TDM, compared with conventional plasma sampling. This study evaluated the feasibility of using the DBS method for Bu TDM. The hematocrit (Hct) and conditioning day were also examined for their impact on the DBS method's performance. Venous blood collected from 6 healthy volunteers was diluted, using their plasma into 4 samples of varying Hct values. Each sample was spiked with Bu calibrators (300, 600, and 1400 ng/mL), prepared using DBS and dried plasma spot (DPS) sampling and analyzed using a validated liquid-chromatography tandem-mass spectrometry method. Clinical blood samples (n = 153) from pediatric patients (n = 15) treated with Bu (mainly from doses 1, 2, 5, and 9) were used to prepare paired volumetric DBS and DPS samples. A Bland-Altman plot and Deming regression were used to define the agreement between the paired DBS and DPS measurements. Passing-Bablok regression analyses investigated the effects of Hct and conditioning day on the linearity between both methods. In vitro analyses showed good agreement between DBS and DPS measurements, with a mean difference of -5.4% and a 95% confidence interval on the limits of agreement of -15.3% to 4.6%. Clinical samples showed good correlation (Pearson correlation coefficient = 0.96; slope = 1.00) between the DBS and DPS methods. The DBS method met the clinical acceptance limits for clinical samples, with a bias <±20%. Bland-Altman plots showed good agreement, with only 5.8% of paired measurements exceeding the limits of agreement (±1.96 SD), although within its 95% confidence interval. Hct observations ranged from 21.7% to 34.7% and did not affect Bu concentrations measured from DBS in either the in vitro or in vivo studies. These results show that DBS is a useful method for Bu TDM, provided samples are analyzed on the collection day. DBS sampling offers advantages over traditional plasma sampling in infants and younger children because only small volumes of blood are required.

Comparative evaluation of urinary dipstick and pH-meter for cattle urine pH measurement

Urinary dipsticks are often used for the diagnosis of kidney, liver, metabolic, and urogenital diseases in cattle clinical practice for the simplicity of use and ease of access. The aims of the present study were to evaluate the agreement between urinary dipsticks and pH-meter for the urine pH in dairy cattle, and also to compare the urine pH before and after centrifugation from each method. The agreement between urine dipstick and pH-meter methods before and after centrifuge were calculated by Passing-Bablok regression analysis demonstrated constant differences (intercept) at 0.60 and -1.01 and positive proportional differences (slope) at 0.94 and 1.13; respectively. Total bias estimated by Bland-Altman plot analysis before (0.20) and after (0.14) centrifuge were lower than the acceptable bias in urine samples. The regression analysis of this study emphasized that the urinary dipstick can be used to determine the cattle urine.

EVALUATION OF 25-HYDROXYVITAMIN D IN HOFFMANN'S TWO-TOED SLOTHS (CHOLOEPUS HOFFMANNI) USING DRIED BLOOD SPOTS ANALYZED BY LIQUID CHROMATOGRAPHY-TANDEM MASS SPECTROMETRY.

Although biochemical analytes have typically been measured using serum or whole blood samples, an increasing number of assays are validated for measurement of analytes from dried blood spots (DBS) on filter paper. DBS techniques are minimally invasive, require only a small sample volume, and simplify processing, storage, and shipment of samples. These qualities make DBS-based assays ideal for sampling of wildlife species in both captive and field settings. In this study, a liquid chromatography-tandem mass spectrometry assay was evaluated for measurement of 25-hydroxyvitamin D in sloths. Paired serum and DBS samples were collected from nine healthy captive Hoffmann's two-toed sloths (Choloepus hoffmanni). Statistical analysis using Passing-Bablok regression analysis, Bland-Altman plots, and the Wilcoxon signed-rank tests found good agreement between 25-hydroxyvitamin D3 measurements in serum and DBS samples. Constant and proportional bias were absent. Results from this study support the use of DBS samples for the evaluation of vitamin D status in Hoffmann's two-toed sloths and provide a foundation for further studies to validate this technique.

Evaluation of the Analytical Performance of Lifotronic H9 Glycated Hemoglobin Analyzer and its Effect on Clinical Decision Making

We aimed to analyze the analytical performance of Lifotronic H9 glycated hemoglobin analyzer (Lifotronic H9) with Arkray Adams HA-8160 glycated hemoglobin analyzer (Arkray HA8160), which is currently used for measurement of HbA1c test in our laboratory. The verification studies of the Lifotronic H9 were performed by comparison with the Arkray HA8160, which has been verified and is already used in our laboratory, and in accordance with the CLSI EP5-A3 and CLSI 15-A3 guidelines. The method comparison study was performed with 316 subjects. Both instruments demonstrated perfect repeatability (within-run) CVs (0.8% - 1.6%), between-run CVs (0.2% - 1.0%), within-day CVs (0.8% - 1.7%), between-day CVs (0.3% - 0.6%), and within laboratory (total im-precision) CVs (0.8% - 1.8%) with two levels of quality control materials. Very good linearity was observed in the method-comparison studies based on Passing-Bablok regression analysis (y = -0.03 + 1.02 x, CUSUM test p = 0.07). Intraclass correlation coefficient (ICC = 0.995) for results and Kappa value (κ = 0.86) for classification of results according to the American Diabetes Association recommendations showed almost perfect agreement. The Lifotronic H9 analyzer shows good precision with lower than recommended total CV 2% and linearity with Arkray HA8160. The Lifotronic H9 is a suitable analyzer for the diagnosis of diabetes and monitoring diabetic control.

Circulating Sex Steroid Measurements of Men by Mass Spectrometry Are Highly Reproducible after Prolonged Frozen Storage

Long-term studies investigating hormone-dependent cancers and reproductive health often require prolonged frozen storage of serum which assumes that the steroid molecules and measurements are stable over that time. Previous studies of reproducibility of circulating steroids have relied upon flawed historical rather than contemporaneous controls. We measured serum testosterone (T), dihydrotestosterone (DHT), estradiol (E2) and estrone (E1) in 150 randomly selected serum samples by liquid chromatography-mass spectrometry (LC-MS) from men 70 years or older (mean age 77 years) in the CHAMP study. The original measurements in 2009 were repeated 10 years later using the identical serum aliquot (having undergone 2-4 freeze-thaw cycles in the interim) in 2019 together with another never-thawed aliquot of the same serum sample. The results of all three sets of measurements were evaluated by Passing-Bablok regression and Bland-Altman difference analysis. Serum androgens (T, DHT) and estrogens (E2, E1) measured by LC-MS display excellent reproducibility when stored for 10 years at -80 C without thawing. Serum T and DHT displayed high level of reproducibility across all three sets of measurements. Multiple freeze-thaw cycles over those storage conditions do not significantly affect serum T, DHT and E1 concentrations but produce a modest increase (21%) in serum E2 measurements.

Evaluation of sxtA and rDNA qPCR assays through monitoring of an inshore bloom of Alexandrium catenella Group 1

Alexandrium catenella (formerly A. tamarense Group 1, or A. fundyense) is the leading cause of Paralytic Shellfish Poisoning in North and South America, Europe, Africa, Australia and Asia. The quantification of A.catenella via sxtA, a gene involved in Paralytic Shellfish Toxin synthesis, may be a promising approach, but has not been evaluated in situ on blooms of A. catenella, in which cell abundances may vary from not detectable to in the order of 106 cells L−1. In this study, we compared sxtA assay performance to a qPCR assay targeted to a species-specific region of ribosomal DNA (rDNA) and an established fluorescent in situ hybridization (FISH) microscopy method. Passing-Bablok regression analyses revealed the sxtA assay to overestimate abundances when <5 cell equivalents A. catenella DNA were analysed, but otherwise was closer to microscopy estimates than the rDNA assay, which overestimated abundance across the full range of concentrations analysed, indicative of a copy number difference between the bloom population and a culture used for assay calibration a priori. In contrast, the sxtA assay performed more consistently, indicating less copy number variation. The sxtA assay was generally reliable, fast and effective in quantifying A. catenella and was predictive of PST contamination of shellfish.

P855Evaluation of epicardial coronary resistance using computed tomography angiography

Abstract Background A Fractional flow reserve (FFR) pullback allows assessing the distribution of pressure loss along the vessel. FFR derived from CT (FFRCT) provides a virtual pullback curve that may also aid in the assessment of epicardial coronary resistance in the non-invasive setting. Purpose The present study aims to determine the accuracy of the virtual FFRCT pullback curve using a motorized invasive FFR pullback as reference in patients with stable coronary artery disease. Methods This is a single centre, prospective study of patients with stable coronary artery disease in whom FFRCT was performed as standard of care for non-invasive assessment. Patients referred to coronary angiography with clinically indicated invasive FFR measurement were included. FFRCT and invasive FFR values were extracted from coronary vessels every 1 mm to generate pullback curves. Invasive FFR pullbacks were acquired using a dedicated device at a speed of 1 mm/s. The area under the pullback curve (AUPC), defined as the sum of areas under the FFR pullback curve, was compared between FFRCT and invasive FFR pullbacks. Lesions were defined based on invasive angiography. FFR gradients in lesions and non-obstructive segments were defined as the difference between FFR values at the proximal and distal edge of the segments. FFR vessel gradient was defined as the difference between the most distal FFR value and the FFR at the ostium of the vessel. Mixed effect model was used to account for the correlation of FFR values within vessels. The agreement between FFRCT and FFR gradients was assessed using the Passing Bablok regression analysis and Bland-Altman methods at the vessel, lesion and non-obstructive level. Results A total of 3172 matched FFRCT and FFR values were obtained in 24 vessels. The correlation coefficient between FFRCT and FFR was 0.76 (95% CI 0.75 to 0.78; p&lt;0.001). The mean difference between the FFRCT and invasive FFR pullback values was 0.07 (LOA −0.11 to 0.24). AUPC was similar between FFRCT and invasive FFR (79.0±16.1 vs. 85.3±16.4, p=0.097); the mean slope of FFRCT pullback curve was steeper compared to invasive FFR (p&lt;0.001). The mean difference in lesion gradient was −0.07 (LOA −0.26 to 0.13) and −0.01 (LOA −0.06 to 0.05) in non-obstructive segments. There were no systematic or proportional differences between FFRCT and FFR gradients either in lesion or non-obstructive segments); however, vessel gradients were overestimated by FFRCT with a bias of −0.12 (LOA −0.35 to 0.12) driven by a higher mean difference in lesion gradients (−0.07; 95% CI −0.26 to 0.13). Conclusions The evaluation of epicardial coronary resistance using coronary CT angiography with FFRCT was feasible. FFRCT pullbacks were accurate in the assessment of lesion and non-obstructive gradients. FFRCT can identify the physiological pattern of coronary artery disease in the non-invasive setting.

Performance Evaluation of a New Mobile POC Lipids Analyzer

Hyperlipidemia, excessive amounts of fat and fatty substances in the blood, is a major risk factor for cardiovascular disease. The measuring of fat or fatty substance in blood is usually obtained through a blood lipid test which includes total cholesterol (TC), triglycerides (TG), low-density lipoprotein (LDL) and high-density lipoprotein (HDL). In the Medical Device industry, Point-of-care (POC) methods provide fast, portable and easy to use devices suitable for rapid determination of Lipid levels. Here, we evaluated the analytical performance of PixoTest POCT Lipid System (iXensor Co., Ltd.), a newly developed POC lipid panel analyzer. PixoTest POCT Analyzer accuracy evaluation with the Cobas c111 Analyzer (Roche Diagnostics Ltd.) as a reference method was carried out according to CLSI EP 09-A3 guidelines. This study enrolled one hundred and twenty (120) subjects and conducted over three (3) clinical sites (40 subjects each), administered through nine (9) trained operators (total) performed under manufacturer’s protocol using three (3) reagent lots per blood sample (capillary & venous). Linear regression, bias plot and other statistical analyses were extrapolated from the test data in this study. Passing-Bablok regression analysis of the data between PixoTest POCT System and Roche Cobas c111 Analyzer with the venous blood showed a slope of 1.03, an intercept of − 0.1 and correlation coefficient of 0.99. The results for capillary blood showed a slope of 1.01, an intercept of − 0.04 and correlation coefficient of 0.99. Passing-Bablok regression analysis of the data (n = 120) from the comparison between the PixoTest POCT System and Roche Cobas c111 Analyzer with the TC venous blood of total 3 lots at 3 sites showed a slope of 0.99, an intercept of − 0.337 and correlation coefficient of 0.995 and the TC capillary blood showed a slope of 0.99, an intercept of − 0.105 and correlation coefficient of 0.984. TG venous blood of total 3 lots at 3 sites showed a slope of 0.99, an intercept of 0.0246 and correlation coefficient of 0.998 and TG capillary blood showed a slope of 0.99, an intercept of − 0.329 and correlation coefficient of 0.998. HDL venous blood showed a slope of 0.99, an intercept of − 0.0297 and correlation coefficient of 0.987 and the HDL capillary blood showed a slope of 0.99, an intercept of − 0.0403 and correlation coefficient of 0.989. The evaluation results of PixoTest POCT system’s accuracy, linearity, and correlation with Roche Cobas c111 Analyzer, the PixoTest POCT System displayed great performance results for all evaluated data points. Thus, the PixoTest POCT System is capable of providing accurate and reliable lipid measurements in the POC setting.

Assessment of Abbott Architect 25-OH vitamin D assay in different levels of vitamin D.

There is increasing requests of Vitamin D test in many clinical settings in recent years. However, immunoassay performance is still a controversial topic. Several diagnostic manufacturers have launched automated 25-hydroxyvitamin D (25-OH D) immunoassays in the past decade. We compared the performance of Abbott Architect 25-OH D Vitamin immunoassay with liquid chromatography-tandem mass spectrometry systems (LCMS/MS) to evaluate immunoassay performance, especially in deficient groups. Eighty human serum samples were analyzed with Architect 25-OH D vitamin kit (Abbott Diagnostics, Lake Forest, IL, USA) and LC-MS/MS systems (Zivak Technology, Istanbul, Turkey). The results of the immunoassay method were compared with the LC-MS/MS using Passing-Bablok regression analysis, Bland-Altman plots and correlation coefficient analysis. We also evaluated results in four levels of D vitamin as a severe deficiency, deficiency, insufficiency, and sufficiency. Architect showed 9.59% bias from LC-MS/MS with smaller mean. Passing-Bablok regression analysis demonstrated the value of 0.95 slope and had a constant bias with an intercept value of -4.25. Concordance correlation coefficient showed moderate agreement with the value of 0.918 (95% CI 0.878-0.945). Two methods revealed good interrater agreement (kappa = 0.738). While the smallest bias determined in deficiency (9.95%) group, the biggest was in insufficiency (15.15%). Architect 25-OH D vitamin immunoassay can be used in routine measurements but had potential misclassification of vitamin D status in insufficient and deficient groups. Although there are recent standardization attempts in 25-OH D measurements, clinical laboratories must be aware of this method.

Advantages of the Use of Alcor iSED Automated Sed Rate Analyzer vs Manual Sed Rate in Evaluation and/or Monitoring the Treatment in Patients, CMCVAMC Experience

Abstract Objectives The automated Alcor iSED automated instrument has a major impact on the proper management of the patients in a timely manner with use of a minimal amount of the blood (100 µL). The Alcor iSED analyzer has introduced a new technology that forgoes sample preparation by using a Load and Go technology. We were previously using the Westergren tube method, which required special preparation of the tube to load into a special provided sealed cup. The entire procedure was tedious and time-consuming. To overcome, we acquired and implemented the Alcor iSED analyzer for an efficient and prompt reporting of sedimentation rate. Methods A sample was aspirated and dispensed from a closed tube, then dispensed into a capillary, and quantitative capillary photometry (aggregation) was used to measure ESR faster than traditional methods by capturing the kinetics of red blood cell aggregation in a controlled testing environment during the most critical phase of sedimentation, commonly referred to as the lag or Rouleaux formation phase. As a part of quality assurance monitoring, retrospective split analysis was performed for 75 samples. Analysis was performed using a Passing-Bablok regression analysis with the MedCalc program. Results Our results demonstrated significant differences between the two methods for the TAT. The TAT was significantly decreased with Alcor iSED (3 minutes for the first specimen and then 15-20 seconds for each sequential specimen) as compared to the manual Westergren method (1 hour), which significantly improved the reporting of results to the providers to monitor the treatment. Conclusion The Alcor iSED analyzer has provided truly shortened TAT erythrocyte sedimentation rate, allowing for better patient care and treatment.

Mobile Health for Diagnosis-PixoTest HbA1c POCT System

Glycated hemoglobin (HbA1c) has been recommended as a standard indicator of long-term glycemic control for diabetic and non-diabetic patients, within the in vitro diagnostic (IVD) industry that is focused on the development of point-of-care (POC) methods with improved operational convenience and shorter measurement time. In this study, we evaluated the measurement accuracy performance of a recently commercialized POC HbA1c analyzer, PixoTest POCT System (iXensor Co., Ltd.) that is portable, intuitive and wireless data transmission enabled. For PixoTest POCT accuracy evaluation, TOSOH G7 Analyzer (TOSOH Bioscience GmbH) was used as a reference method and performed in accordance with CLSI EP 09-A3 guidelines. This study enrolled one hundred and twenty (120) subjects from three clinical sites (40 subjects each), administered through nine (9) trained operators (total) performed under manufacturer’s protocol using three reagent lots per blood sample (capillary & venous). Linear regression, bias plot and other statistical analyses were extrapolated from the test data. Passing-Bablok regression analysis of the data between PixoTest POCT System and TOSOH G7 Analyzer with the venous blood showed a slope of 1.03 and an intercept of − 0.1 and a correlation coefficient of 0.99. The results for capillary blood showed a slope of 1.01 and an intercept of − 0.04 and a correlation coefficient of 0.99. The PixoTest POCT System’s performance is similar to TOSOH G7 Analyzer in accuracy and linearity. Therefore, the system’s HbA1c test capability is ideal for the management of chronic disease, as a user convenient and efficient POC solution.

Toward a Harmonized and Standardized Protocol for the Determination of Total Hydroxytyrosol and Tyrosol Content in Virgin Olive Oil (VOO). The Pros of a Fit for the Purpose Ultra High Performance Liquid Chromatography (UHPLC) Procedure.

Τoward a harmonized and standardized procedure for the determination of total hydroxytyrosol and tyrosol content in virgin olive oil (VOO), the pros of a recently published in house validated ultra high performance liquid chromatography (UHPLC) protocol are discussed comparatively with those of other procedures that determine directly or indirectly the compounds hosted under the health claim on “olive oil polyphenols” (EC regulation 432/2012). Authentic VOOs were analyzed with five different liquid chromatographic separation protocols and 1H-NMR one in five different laboratories with expertise in VOO phenol analysis within three months. Data comparison indicated differences in absolute values. Method comparison using appropriate tools (Passing-Bablok regression and Bland Altman analyses) for all protocols vs. the UHPLC one indicated slight or statistically significant differences. The results were also discussed in terms of cost effectiveness, detection means, standard requirements and ways to calculate the total hydroxytyrosol and tyrosol content. Findings point out that the in-house validated fit for the purpose UHPLC protocol presents certain pros that should be exploited by the interested parties. These are the simplicity of sample preparation, fast elution time that increase the number of samples analyzed per day and integration of well-resolved peaks with the aid of only two commercially available external standards. Importance of correction factors in the calculations is stressed.

Heavy/light chain assay in the monitoring of multiple myeloma

Serum protein (SPE) and immunofixation electrophoresis (IFE) have been extensively validated for the routine use of identifying, characterising and quantifying monoclonal proteins. However, accurate quantitation of IgA monoclonal proteins can be difficult when they migrate in to the β fraction, due to co-migration with transferrin and complement components. The heavy/light chain (HLC) immunoassay is an additional tool for measuring intact immunoglobulin monoclonal proteins. Therefore, we aimed to examine the clinical utility of the HLC assay for the disease monitoring of IgG and IgA multiple myeloma (MM) patients. A total of 177 samples from 30 MM patients (21 IgG and 9 IgA) were analysed retrospectively with median number of six follow up samples per patient (range 3-13). Serum free light chains (sFLC) and HLC were quantified using Freelite and Hevylite immunoassays. Details of M-protein concentration, β-globulin levels, total immunoglobulin levels and disease treatment response were obtained from the laboratory and patient information system. Passing-Bablok regression analysis was performed to compare (i) M-protein quantification with involved HLC (iHLC) and (ii) total immunoglobulin with summated HLC pairs for each immunoglobulin type (e.g., IgGκ+IgGλ). For 127 IgG MM samples, IgG iHLC levels showed a good correlation with SPE quantification (iHLC y=0.96x+4.9; r=0.917) and summated HLC showed a good correlation with total IgG concentration (summated HLC y=0.94x+5.74; r=0.91). In total, 95/127 (75%) IgG MM follow-up samples had an abnormal HLC ratio and 122/127 (96%) had a positive SPE, probably due to the lower sensitivity of HLC assay in detecting clonality in patients with IgG MM. Consistent with this, one patient assigned a very good partial response by International Myeloma Working Group criteria would be assigned a complete response based on HLC measurements. For 50 IgA MM samples, 42/50 (84%) had an abnormal HLC ratio. Conversely, 50/50 (100%) of M-proteins showed β fraction migration and were difficult to accurately quantify by SPE. Therefore, M-protein concentration and iHLC did not correlate as well in IgA MM (y=1.9x-8.4; r=0.8) compared to IgG MM. However, there was good correlation between total IgA and summated IgA HLC (IgAκ+IgAλ y=1.35x-0.33; r=0.95). Of the 8/50 (16%) IgA samples with a normal HLC ratio, 6/8 (75%) were consistent with the disease status being in complete remission. Interestingly, in one IgA MM patient, SPE and IFE were negative, but the serum FLC ratio and involved FLC were highly abnormal, consistent with the presence of light chain escape. Our data suggest HLC measurements could add value to the current disease monitoring of MM patients. In IgG MM patients, the M-protein level correlated well with HLC values. The HLC assay complements the serum FLC assay and is especially useful for monitoring of IgA MM patients who display M-proteins migrating in the β region on SPE.

Analytical comparison between two hematological analyzer systems: Mindray BC‐5180 vs Sysmex XN‐1000

BackgroundTo compare the Mindray BC‐5180 and Sysmex XN‐1000 instruments by analyzing the results of complete blood count in the external quality assessment in Shandong Province in 2018.MethodsIn the external quality assessment, 10 batches of quality control materials were issued throughout the year. The test items were WBC, RBC, Hb, PLT, and HCT. The laboratories using Mindray BC‐5180 and Sysmex XN‐1000 were screened, and the results were analyzed by t test, Passing‐Bablok regression analysis, and Bland‐Altman analysis.ResultsThirty‐six laboratories using Mindray BC‐5180 instruments and thirty‐six laboratories using Sysmex XN‐1000 instruments were screened, and the average difference between the two instruments results is not significant (P > 0.05, t test). Passing‐Bablok regression analysis showed that the 95% confidence interval of the regression equation interception of each test item included 0, and the 95% confidence interval of the slope contained 1, r > 0.98, which showed that the correlation is good. The Bland‐Altman analysis showed that both instruments had more than 95% of the points within the 95% consistency limit (WBC97.2%, RBC95.6%, PLT97.2%, Hb96.7%, HCT97.5%). Within the consistency limit, the absolute value of the difference between the Mindray BC‐5180 instrument and the Sysmex XN‐1000 instrument is WBC 0.14%, RBC 0.26%, PLT 2.7%, and Hb 1.9%. HCT is 0.69%, and the difference is clinically acceptable.ConclusionIt can be considered that the two instruments have good correlation and consistency, and the two instruments can replace each other.

In vitro validation of indirect calorimetry device developed for the ICALIC project against mass spectrometry

Accurate evaluation of the energy needs is required to optimize nutrition support of critically ill patients. Recent evaluations of indirect calorimeters revealed significant differences among the devices available on the market. A new indirect calorimeter (Q-NRG®, Cosmed, Roma, Italy) has been developed by a group of investigators supporting the international calorimetry study initiative (ICALIC) to achieve ultimate accuracy for measuring energy expenditure while being easy to use, and affordable. This study aims to validate the precision and the accuracy of the Q-NRG® in the in-vitro setting, within the clinically relevant range for adults on mechanical ventilation in the ICU. Mass spectrometry is the reference method for the gas composition analysis to evaluate the analytic performances of the Q-NRG®. The accuracy and precision of the O2 and CO2 measurements by the Q-NRG were evaluated by comparing the measurements of known O2 and CO2 gas mixtures with the measurements by the mass spectrometer (Extrel, USA). The accuracy and precision of the Q-NRG® for measurements of VO2 (oxygen consumption) and VCO2 (CO2 production) at clinically relevant ranges (150, 250 and 400ml/min STPD) were evaluated by measuring simulated gas exchange under mechanically ventilated setting at different FiO2 settings (21-80%), in comparison to the reference measurements by the mass spectrometer-based mixing chamber system. The measurements of gas mixtures of predefined O2 and CO2 concentrations by the Q-NRG® were within 2% accuracy versus the mass spectrometer measurements in Passing Bablok regression analysis. In a mechanically ventilated setting of FiO2 from 21 up to 70%, the Q-NRG® measurements of simulated VO2 and VCO2 were within 5% difference of the reference mass spectrometer measurements. Invitro evaluation confirms that the accuracy of the Q-NRG® indirect calorimeter is within 5% at oxygen enrichment to 70%; i.e. maximum expected for clinical use. Further recommendations for the clinical use of the Q-NRG® by will be released once the ongoing multi-center study is completed.

QIP-MS: A specific, sensitive, accurate, and quantitative alternative to electrophoresis that can identify endogenous m-proteins and distinguish them from therapeutic monoclonal antibodies in patients being treated for multiple myeloma

Serum protein (SPE) and immunofixation electrophoresis (IFE) have been extensively validated for the routine use of identifying, characterising and quantifying monoclonal proteins. However, accurate quantitation of IgA monoclonal proteins can be difficult when they migrate in to the β fraction, due to co-migration with transferrin and complement components. The heavy/light chain (HLC) immunoassay is an additional tool for measuring intact immunoglobulin monoclonal proteins. Therefore, we aimed to examine the clinical utility of the HLC assay for the disease monitoring of IgG and IgA multiple myeloma (MM) patients.A total of 177 samples from 30 MM patients (21 IgG and 9 IgA) were analysed retrospectively with median number of six follow up samples per patient (range 3–13). Serum free light chains (sFLC) and HLC were quantified using Freelite and Hevylite immunoassays. Details of M-protein concentration, β-globulin levels, total immunoglobulin levels and disease treatment response were obtained from the laboratory and patient information system. Passing–Bablok regression analysis was performed to compare (i) M-protein quantification with involved HLC (iHLC) and (ii) total immunoglobulin with summated HLC pairs for each immunoglobulin type (e.g., IgGκ+IgGλ).For 127 IgG MM samples, IgG iHLC levels showed a good correlation with SPE quantification (iHLC y=0.96x+4.9; r=0.917) and summated HLC showed a good correlation with total IgG concentration (summated HLC y=0.94x+5.74; r=0.91). In total, 95/127 (75%) IgG MM follow-up samples had an abnormal HLC ratio and 122/127 (96%) had a positive SPE, probably due to the lower sensitivity of HLC assay in detecting clonality in patients with IgG MM. Consistent with this, one patient assigned a very good partial response by International Myeloma Working Group criteria would be assigned a complete response based on HLC measurements. For 50 IgA MM samples, 42/50 (84%) had an abnormal HLC ratio. Conversely, 50/50 (100%) of M-proteins showed β fraction migration and were difficult to accurately quantify by SPE. Therefore, M-protein concentration and iHLC did not correlate as well in IgA MM (y=1.9x–8.4; r=0.8) compared to IgG MM. However, there was good correlation between total IgA and summated IgA HLC (IgAκ+IgAλ y=1.35x–0.33; r=0.95). Of the 8/50 (16%) IgA samples with a normal HLC ratio, 6/8 (75%) were consistent with the disease status being in complete remission. Interestingly, in one IgA MM patient, SPE and IFE were negative, but the serum FLC ratio and involved FLC were highly abnormal, consistent with the presence of light chain escape.Our data suggest HLC measurements could add value to the current disease monitoring of MM patients. In IgG MM patients, the M-protein level correlated well with HLC values. The HLC assay complements the serum FLC assay and is especially useful for monitoring of IgA MM patients who display M-proteins migrating in the β region on SPE.

Comparability and precision of four oncological methods on seven immunochemistry analysers

Siemens Healthcare Diagnostics has four commercially available assays on different analytical platforms using different methodologies to generate signal. We assessed the analytical performance of the Dimension EXL hs-cTnI assay (LOCI method) across different matrices and compared it to two different acridinium ester-based hs-cTnI assays (ADVIA Centaur and Abbott ARCHITECT).The analytical sensitivity and precision below the 99th-percentile was determined for the Dimension EXL hs-cTnI assay. Method comparisons were performed between the Dimension EXL contemporary cTnI and the hs-cTnI assays, between different matrices for the EXL hs-cTnI assay (serum, lithium heparin and EDTA plasma), and between different hs-cTnI assays (EXL versus ADVIA Centaur or Abbott ARCHITECT) using non-parametric analyses.The limit of blank and detection were 0.9 ng/L and 1.7 ng/L, respectively, with imprecision of 5.8% at 8.6 ng/L and 3.2% at 47.5 ng/L. Comparison between the EXL contemporary cTnI and hs-cTnI assay (range: 2.6–4214 ng/L) yielded proportional lower concentrations for the hs-cTnI assay (slope = 0.86; 95%CI: 0.81 to 0.96, n = 40); however, there was no difference in concentrations below 100 ng/L between the assays (median difference = −2.7 ng/L; 95%CI: −9.8 to 9.3). Passing-Bablok regression analysis with EDTA plasma yielded proportionally higher concentrations with the EXL hs-cTnI versus Abbott hs-cTnI (slope = 1.45; 95%CI: 1.02–1.86, n = 40) with proportionally lower concentrations with EDTA versus lithium heparin plasma with the EXL hs-cTnI assay alone (slope = 0.93; 95%CI: 0.90 to 0.99, n = 40). Comparison with Abbott hs-cTnI concentrations below 100 ng/L in the three matrices, indicated that the EXL hs-cTnI assay yielded higher concentrations (median difference range: 3.4–9.4 ng/L), with differences also evident when comparing the EXL hs-cTnI assay to the ADVIA Centaur hs-cTnI assay.The Siemens EXL hs-cTnI assay meets the analytical criteria for a high-sensitivity assay, with assay specific cutoffs important to maximize clinical performance.

Determination of sigma score based on biological variation for haemostasis assays: Fit-for-purpose for daily practice?

The Abbott Alinity family of chemistry and immunoassay systems recently launched with early adopters contributing imprecision and bias data, which was consolidated to assess the performance of Alinity assays across multiple sites using the Sigma metric. Multi-site Sigma metrics were determined for 3 ion-selective electrodes, 12 photometric assays, and 3 immunoassays across 11 independent laboratory sites in 9 countries.Total allowable error (TEa) goals followed a previously defined hierarchy that used CLIA as the primary goal. Bias was calculated against the Abbott ARCHITECT system using Passing-Bablok regression analysis using individual site data or pooled aggregate data. Sigma metrics were calculated as (%TEa - |% bias|)/%CV. For individual-site analysis, the Sigma metrics for each assay were compared using the individual-site and the pooled biases. For multi-site analysis, the average CV and the pooled bias were used to generate a Pooled Sigma metric encompassing the global performance for a given assay.A total of 97 individual-site and 18 Pooled Sigma metrics were calculated for available assays. Individual Sigma metrics varied across sites, with 90% of assays performing 4 Sigma or higher, and 17 of 18 Pooled Sigma metrics indicated performance greater than 4 Sigma. Sigma metrics were significantly improved in 16 assays when using pooled bias rather than individual-site bias.This multi-center study applies a novel application of Sigma metrics to the first Alinity users and reveals analytical performance of greater than 4 Sigma for vast majority of assays. Laboratories with limited resources can leverage larger data sets for Pooled Sigma metric analysis, providing a tool to assess the consistency of analytical performance from multiple sites.

Analytical characterization of the Siemens Dimension EXL high-sensitivity cardiac troponin I assay

BackgroundSiemens Healthcare Diagnostics has four commercially available assays on different analytical platforms using different methodologies to generate signal. We assessed the analytical performance of the Dimension EXL hs-cTnI assay (LOCI method) across different matrices and compared it to two different acridinium ester-based hs-cTnI assays (ADVIA Centaur and Abbott ARCHITECT). MethodsThe analytical sensitivity and precision below the 99th-percentile was determined for the Dimension EXL hs-cTnI assay. Method comparisons were performed between the Dimension EXL contemporary cTnI and the hs-cTnI assays, between different matrices for the EXL hs-cTnI assay (serum, lithium heparin and EDTA plasma), and between different hs-cTnI assays (EXL versus ADVIA Centaur or Abbott ARCHITECT) using non-parametric analyses. ResultsThe limit of blank and detection were 0.9 ng/L and 1.7 ng/L, respectively, with imprecision of 5.8% at 8.6 ng/L and 3.2% at 47.5 ng/L. Comparison between the EXL contemporary cTnI and hs-cTnI assay (range: 2.6–4214 ng/L) yielded proportional lower concentrations for the hs-cTnI assay (slope = 0.86; 95%CI: 0.81 to 0.96, n = 40); however, there was no difference in concentrations below 100 ng/L between the assays (median difference = −2.7 ng/L; 95%CI: −9.8 to 9.3). Passing-Bablok regression analysis with EDTA plasma yielded proportionally higher concentrations with the EXL hs-cTnI versus Abbott hs-cTnI (slope = 1.45; 95%CI: 1.02–1.86, n = 40) with proportionally lower concentrations with EDTA versus lithium heparin plasma with the EXL hs-cTnI assay alone (slope = 0.93; 95%CI: 0.90 to 0.99, n = 40). Comparison with Abbott hs-cTnI concentrations below 100 ng/L in the three matrices, indicated that the EXL hs-cTnI assay yielded higher concentrations (median difference range: 3.4–9.4 ng/L), with differences also evident when comparing the EXL hs-cTnI assay to the ADVIA Centaur hs-cTnI assay. ConclusionThe Siemens EXL hs-cTnI assay meets the analytical criteria for a high-sensitivity assay, with assay specific cutoffs important to maximize clinical performance.

The pathway for introducing novel examination procedures in routine practice in accordance with ISO 15189:2012: 17-Hydroxy progesterone, dehydroepiandrosterone sulphate and vitamin D as examples.

Background In a medical laboratory, changes may be made to the analytical phase of diagnostic testing whenever a new test or the issue of a ‘new generation’ kit or new diagnostic system is required. In such cases, ISO 15189:2012 accreditation can assist laboratory professionals. The aim of the present study was to propose a working pathway for introducing new examination procedures into clinical practice in accordance with the ISO 15189:2012 standard, through the exemplars of 17-hydroxy progesterone, dehydroepiandrosterone sulphate and vitamin D. Methods The working pathway includes the following steps: (i) analysing examination procedures under evaluation, (ii) analysing examination procedures currently in use, (iii) verifying metrological traceability, (iv) verifying examination procedures and (v) evaluating comparability of results. Results The analysis of instructions for use issued by manufacturers revealed that metrological traceability was reported only for vitamin D. The imprecision verification was satisfactory, the imprecision obtained by the laboratory in terms of total imprecision always being less than the specified total imprecision. In only one case (IQC level 1, 17-hydroxy progesterone), the total upper verification limit was calculated. The trueness verification was satisfactory for all examination procedures, except for 17-hydroxy progesterone (second material). Passing–Bablok regression analyses in the comparability study demonstrated significant differences for all the examination procedures. Conclusions The working pathway described for examination procedures in routine practice is in accordance with the requirements of ISO 15189:2012 accreditation and takes feasibility into account (as its main goal), based on the cost/patient benefit ratio.