Effects of different tube types on patient classification using current diabetes decision limits.

Management of diabetes is a challenge starting in the pre-analytical phase with selecting the most appropriate glycolysis inhibitor. Study goal was to calculate the impact of tubes with different glycolysis inhibitors on the classification of the glycemic control of 157,415 consecutive hospital patients according to current WHO diabetes criteria. METHODS: Glucose and lactate were measured in parallel in samples from 68 healthy subjects collected and stored in different sample tubes from Sarstedt and Greiner. Bias to baseline conditions (fluoride heparin (FH) tubes, centrifugation within 1 h) was determined. RESULTS: In baseline samples, glucose concentration in fluoride/EDTA/citrate (FC) plasma was ~13% higher and lactate concentration ~20% lower compared to FH, fluoride oxalate, and fluoride EDTA plasma, and in serum. Glucose recovery after storage up to 48 h was 99-101% in the different tubes, but the effectiveness of glycolysis inhibition by FC was inconsistent. Based on the observed mean bias of 12% when FC tubes are used, we estimate an increase of 48.4-55.8% in the frequency of patients with impaired glucose levels using current WHO criteria. CONCLUSION: Using current established decision limits, the number of patients with impaired glucose levels in the hospital would increase substantially with a strong impact on patient treatment and consumption of resources. The unpredictable failure of glycolysis inhibition in FC tubes does not allow to adjust the decision limits by a fixed factor. In the absence of prospective outcome studies with FC tubes, we recommend to measure glucose in samples containing FH.

Practical guide for identifying unmet clinical needs for biomarkers.

The development and evaluation of novel biomarkers and testing strategies requires a close examination of existing clinical pathways, including mapping of current pathways and identifying areas of unmet need. This approach enables early recognition of analytical and clinical performance criteria to guide evaluation studies, in a cyclical and iterative manner, all the time keeping the clinical pathway and patient health outcomes as the key drivers in the process. The Test Evaluation Working Group of the European Federation of Clinical Chemistry and Laboratory Medicine (EFLM TE-WG) https://www.eflm.eu/site/page/a/1158 has published a conceptual framework of the test evaluation cycle which is driven by the clinical pathway, inherent to which is the test purpose and role within the pathway that are defined by clinical need. To supplement this framework, the EFLM TE-WG has also published an interactive checklist for identifying unmet clinical needs for new biomarkers; a practical tool that laboratories, clinicians, researchers and industry can equally use in a consistent manner when new tests are developed and before they are released to the market. It is hoped that these practical tools will provide consistent and appropriate terminology in this diverse field and offer a platform that facilitates greater consultation and collaboration between all stakeholders. The checklist should assist the work of all colleagues involved in the discovery of novel biomarkers and implementation of new medical tests. The tool is aligned with the IOM recommendations and the FDA and CE regulating body's requirements.

Impact of lipid markers and high-sensitivity C-reactive protein on the value of the 99th percentile upper reference limit for high-sensitivity cardiac troponin I.

OBJECTIVES: i) To assess the relationship between lipid markers and high-sensitivity C-reactive protein (hs-CRP), and high-sensitivity cardiac troponin I (hs-cTnI) in the reference population, and ii) to evaluate the impact of lipid markers and hs-CRP on the 99th percentile upper reference limit (URL) for hs-cTnI. METHODS: 531 questionnaire-identified presumably healthy individuals were enrolled in a single-center, cross-sectional study. Surrogate biomarkers for diabetes, myocardial and renal dysfunction were used to refine the healthy cohort (n=408). Lipid profile, total cholesterol:high-density lipoprotein cholesterol (HDL-C) ratio, non-HDL-C, apolipoprotein AI (apoAI), apolipoprotein B (apoB), apoB:apoAI ratio, lipoprotein(a), small dense low-density lipoprotein cholesterol (LDL-C) and hs-CRP were determined. RESULTS: Individuals with detectable vs. non-detectable hs-cTnI concentrations more often showed elevated LDL-C (60% vs. 46%; p=0.002), apoB (73% vs. 61%; p=0.008), apoB:apoAI ratio (53% vs. 40%; p=0.005) and lipoprotein(a) (15% vs. 7%; p=0.015). The apoB:apoAI ratio and to a lesser extent other lipid markers, but not hs-CRP, were positively associated with hs-cTnI concentration in univariate and multivariate analyses. Exclusion of individuals with elevated apoB:apoAI ratio or apoB, but not hs-CRP, lowered the 99th percentile URL in the healthy cohort respectively by 12.9% (6.2 vs. 5.4ng/L) and 14.5% (6.2 vs. 5.3ng/L). The corresponding reduction for both lipid biomarkers in the presumably healthy population was 24.0% (7.5 vs. 5.7ng/L). CONCLUSION: Our study demonstrates that atherogenic lipid markers, particularly apoB:apoAI ratio or apoB, influence the 99th percentile URL for hs-cTnI.

Biomarker development targeting unmet clinical needs.

BACKGROUND: The introduction of new biomarkers can lead to inappropriate utilization of tests if they do not fill in existing gaps in clinical care. We aimed to define a strategy and checklist for identifying unmet needs for biomarkers. METHODS: A multidisciplinary working group used a 4-step process: 1/ scoping literature review; 2/ face-to-face meetings to discuss scope, strategy and checklist items; 3/ iterative process of feedback and consensus to develop the checklist; 4/ testing and refinement of checklist items using case scenarios. RESULTS: We used clinical pathway mapping to identify clinical management decisions linking biomarker testing to health outcomes and developed a 14-item checklist organized into 4 domains: 1/ identifying and 2/ verifying the unmet need; 3/ validating the intended use; and 4/ assessing the feasibility of the new biomarker to influence clinical practice and health outcome. We present an outcome-focused approach that can be used by multiple stakeholders for any medical test, irrespective of the purpose and role of testing. CONCLUSIONS: The checklist intends to achieve more efficient biomarker development and translation into practice. We propose the checklist is field tested by stakeholders, and advocate the role of the clinical laboratory professional to foster trans-sector collaboration in this regard.

Setting analytical performance specifications based on outcome studies - is it possible?

The 1st Strategic Conference of the European Federation of Clinical Chemistry and Laboratory Medicine proposed a simplified hierarchy for setting analytical performance specifications (APS). The top two levels of the 1999 Stockholm hierarchy, i.e., evaluation of the effect of analytical performance on clinical outcomes and clinical decisions have been proposed to be replaced by one outcome-based model. This model can be supported by: (1a) direct outcome studies; and (1b) indirect outcome studies investigating the impact of analytical performance of the test on clinical classifications or decisions and thereby on the probability of patient relevant clinical outcomes. This paper reviews the need for outcome-based specifications, the most relevant types of outcomes to be considered, and the challenges and limitations faced when setting outcome-based APS. The methods of Model 1a and b are discussed and examples are provided for how outcome data can be translated to APS using the linked evidence and simulation or decision analytic techniques. Outcome-based APS should primarily reflect the clinical needs of patients; should be tailored to the purpose, role and significance of the test in a well defined clinical pathway; and should be defined at a level that achieves net health benefit for patients at reasonable costs. Whilst it is acknowledged that direct evaluations are difficult and may not be possible for all measurands, all other forms of setting APS should be weighed against that standard, and regarded as approximations. Better definition of the relationship between the analytical performance of tests and health outcomes can be used to set analytical performance criteria that aim to improve the clinical and cost-effectiveness of laboratory tests.

Defining normality in a European multinational cohort: Critical factors influencing the 99th percentile upper reference limit for high sensitivity cardiac troponin I.

OBJECTIVE: To establish and critically evaluate the 99th percentile upper reference limit (URL) for high-sensitivity cardiac troponin I (hs-cTnI) in a large healthy European cohort using different selection criteria. METHODS: 1368 presumably healthy individuals from 9 countries were evaluated with surrogate biomarkers for diabetes (glycated hemoglobin [HbA1c] < 48 mmol/mol), myocardial (B-type natriuretic peptide [BNP] < 35 pg/mL) and renal dysfunction (estimated glomerular filtration rate [eGFR] >60 mL/min/1.73 m(2)), and dyslipidemia to refine the healthy cohort. The 99th percentile URLs were independently determined by the non-parametric and robust methods. RESULTS: The use of biomarker selection criteria resulted in a decrease of the 99th percentile URL for hs-cTnI from 23.7 to 14.1 ng/L and from 11.2 to 7.1 ng/L, when using the non-parametric percentile and robust methods, respectively; a further reduction after exclusion of individuals with dyslipidemia was noted. Male gender, BNP, HbA1c and smoking status were independently associated with hs-cTnI concentration in the presumably healthy population, while the impact of age, present in the univariate analysis, decreased after adjustments for gender and surrogate biomarkers. The BNP-based inclusion criterion had the most pronounced effect on the 99th percentile URL, excluding 21% of the study participants and decreasing its value to 11.0 (7.1) ng/L according to the non-parametric (robust) method. Gender, but not age-specific, differences at 99th percentile URL have been identified. CONCLUSION: The selection of a reference population has a critical impact on the 99th percentile value for hs-cTnI. A uniform protocol for the selection of the healthy reference population is needed.

Evaluation of a next generation direct whole blood enzymatic assay for hemoglobin A1c on the ARCHITECT c8000 chemistry system.

BACKGROUND: The utility of HbA1c for the diagnosis of type 2 diabetes requires an accurate, precise and robust test measurement system. Currently, immunoassay and HPLC are the most popular methods for HbA1c quantification, noting however the limitations associated with some platforms, such as imprecision or interference from common hemoglobin variants. Abbott Diagnostics has introduced a fully automated direct enzymatic method for the quantification of HbA1c from whole blood on the ARCHITECT chemistry system. METHODS: Here we completed a method evaluation of the ARCHITECT HbA1c enzymatic assay for imprecision, accuracy, method comparison, interference from hemoglobin variants and specimen stability. This was completed at three independent clinical laboratories in North America and Europe. RESULTS: The total imprecision ranged from 0.5% to 2.2% CV with low and high level control materials. Around the diagnostic cut-off of 48 mmol/mol, the total imprecision was 0.6% CV. Mean bias using reference samples from IFCC and CAP ranged from -1.1 to 1.0 mmol/mol. The enzymatic assay also showed excellent agreement with HPLC methods, with slopes of 1.01 and correlation coefficients ranging from 0.984 to 0.996 compared to Menarini Adams HA-8160, Bio-Rad Variant II and Variant II Turbo instruments. Finally, no significant effect was observed for erythrocyte sedimentation or interference from common hemoglobin variants in patient samples containing heterozygous HbS, HbC, HbD, HbE, and up to 10% HbF. CONCLUSIONS: The ARCHITECT enzymatic assay for HbA1c is a robust and fully automated method that meets the performance requirements to support the diagnosis of type 2 diabetes.

European multicenter analytical evaluation of the Abbott ARCHITECT STAT high sensitive troponin I immunoassay.

BACKGROUND: International recommendations highlight the superior value of cardiac troponins (cTns) for early diagnosis of myocardial infarction along with analytical requirements of improved precision and detectability. In this multicenter study, we investigated the analytical performance of a new high sensitive cardiac troponin I (hs-cTnI) assay and its 99th percentile upper reference limit (URL). METHODS: Laboratories from nine European countries evaluated the ARCHITECT STAT high sensitive troponin I (hs-TnI) immunoassay on the ARCHITECT i2000SR/i1000SR immunoanalyzers. Imprecision, limit of blank (LoB), limit of detection (LoD), limit of quantitation (LoQ) linearity of dilution, interferences, sample type, method comparisons, and 99th percentile URLs were evaluated in this study. RESULTS: Total imprecision of 3.3%-8.9%, 2.0%-3.5% and 1.5%-5.2% was determined for the low, medium and high controls, respectively. The lowest cTnI concentration corresponding to a total CV of 10% was 5.6 ng/L. Common interferences, sample dilution and carryover did not affect the hs-cTnI results. Slight, but statistically significant, differences with sample type were found. Concordance between the investigated hs-cTnI assay and contemporary cTnI assay at 99th percentile cut-off was found to be 95%. TnI was detectable in 75% and 57% of the apparently healthy population using the lower (1.1 ng/L) and upper (1.9 ng/L) limit of the LoD range provided by the ARCHITECT STAT hs-TnI package insert, respectively. The 99th percentile values were gender dependent. CONCLUSIONS: The new ARCHITECT STAT hs-TnI assay with improved analytical features meets the criteria of high sensitive Tn test and will be a valuable diagnostic tool.

Analytical evaluation of the automated galectin-3 assay on the Abbott ARCHITECT immunoassay instruments.

BACKGROUND: Galectin-3 is secreted from macrophages and binds and activates fibroblasts forming collagen. Tissue fibrosis is central to the progression of chronic heart failure (CHF). We performed a European multicentered evaluation of the analytical performance of the two-step routine and Short Turn-Around-Time (STAT) galectin-3 immunoassay on the ARCHITECT i1000SR, i2000SR, and i4000SR (Abbott Laboratories). METHODS: We evaluated the assay precision and dilution linearity for both routine and STAT assays and compared serum and plasma, and fresh vs. frozen samples. The reference interval and biological variability were also assessed. Measurable samples were compared between ARCHITECT instruments and between the routine and STAT assays and also to a galectin-3 ELISA (BG Medicine). RESULTS: The total assay coefficient of variation (CV%) was 2.3%-6.2% and 1.7%-7.4% for the routine and STAT assays, respectively. Both assays demonstrated linearity up to 120 ng/mL. Galectin-3 concentrations were higher in plasma samples than in serum samples and correlated well between fresh and frozen samples (R=0.997), between the routine and STAT assays, between the ARCHITECT i1000 and i2000 instruments and with the galectin-3 ELISA. The reference interval on 627 apparently healthy individuals (53% male) yielded upper 95th and 97.5th percentiles of 25.2 and 28.4 ng/mL, respectively. Values were significantly lower in subjects younger than 50 years. CONCLUSIONS: The galectin-3 routine and STAT assays on the Abbott ARCHITECT instruments demonstrated good analytical performance. Further clinical studies are required to demonstrate the diagnostic and prognostic potential of this novel marker in patients with CHF.