High-sensitivity Assay Research Articles

SAT-501 Thyrotoxicosis with Pre-Ventricular Complexes Resulting in Thyroid Storm and Cardiac Arrest

Background: Ventricular arrythmias are a rare, often lethal complication of thyrotoxicosis. We describe a patient with uncontrolled hyperthyroidism and pre-ventricular complexes (PVCs) who presented with ventricular tachyarrhythmia cardiac arrest and was successfully resuscitated. Clinical Case: A 64 year old woman was diagnosed with thyrotoxicosis secondary to Graves’ disease [TSH < 0.01 (0.40 – 4.5 mcIU/mL) and free T4 of 2.8 (0.8 – 1.8 ng/dL)] 1 year ago in the setting of a 6 month history of weight loss, palpitations, tremors, and a large goiter. She was started on methimazole and metoprolol XL and was intermittently compliant. During follow-up evaluation she complained of light headedness, developed agonal breathing, and became pulseless. Chest compressions were initiated. She regained spontaneous rhythm after receiving 1 shock with an Automated Electronic Defibrillator (AED). She was transferred to the Emergency Room (ER) and intubated for altered mental status.Emergent CT Angiography and bedside echocardiogram showed no pulmonary embolism and normal biventricular function. Troponin T high sensitivity assay was negative and electrolytes were normal. Repeat thyroid function tests showed TSH <0.01, Free T4 of 5.6 and free T3 of 14.5 (2.0 – 4.4 pg/mL). She was started on propylthiouracil, glucocorticoids, potassium iodide and treated for thyroid storm. EKG in the ER showed sinus tachycardia with no ischemic ST changes but PVCs and fusion complexes were noted. These were also present on EKG at the time of her initial diagnosis of hyperthyroidism. EKGs prior to the diagnosis of hyperthyroidism showed normal sinus rhythm.Cardiac arrest was attributed to thyrotoxicosis as there was no infectious nidus and no evidence of structural cardiac disease. The AED rhythm strips could not be obtained but she was presumed to have an appropriately shockable ventricular tachyarrhythmia such as ventricular tachycardia (VT) or ventricular fibrillation (VF).Her thyroid hormone levels declined appropriately over the course of the hospitalization and PVCs were no longer noted on telemetry and daily EKGs. She was discharged on methimazole which she took consistently. She underwent RAI ablation several months after discharge. Conclusion: Failure to achieve rapid euthyroidism in thyrotoxicosis is associated with increased cardiovascular morbidity and mortality (1). Most arrythmias associated with thyrotoxicosis are supraventricular and ventricular arrythmias are a rare sequela (2). This is one of the few cases reported of antecedent PVCs being noted on EKG. The PVCs resolved with anti-thyroid medications.

MON-111 Plasma Insulin Measured with a Sensitive Immunoassay May Establish the Diagnosis of Congenital Hyperinsulinism Without Further Testing

Background: The diagnosis of congenital hyperinsulinism (CHI) is often hampered by a plasma insulin (p-insulin) detection limit of 2-3 mU/L (14-21 pmol/L) by RIA methods. Objective: To evaluate the diagnostic performance of a sensitive immunoassay for p-insulin and to find the optimal p-insulin cut-off for CHI versus other conditions with hypoglycaemia. Design: Single centre retrospective cohort study. Methods: Diagnostic tests with no medication, no i.v. glucose and under fasting conditions were performed in children with a clinical diagnosis of CHI. P-insulin concentrations determined at simultaneous p-glucose concentrations at least <3.2 mmol/L (57.5 mg/dL) were included in the analysis (n=61).The diagnosis of CHI was either clinical (n=61) or by gold standard criteria: hypoketotic hypoglycaemia plus disease-causing genetic mutations and/or diffuse, focal or atypical pancreatic histopathology (n=57). Samples from 15 children with idiopathic ketotic hypoglycaemia (IKH, diagnosis by exclusion, p-ketones >1.5 mmol/L during hypoglycaemia) were used as controls.P-insulin was measured by the high-sensitive assay (Cobas e411 immunoassay analyzer); lower detection limit 1.4 pmol/L (0.2 mU/L); normal range 18-173 pmol/L (2.57-24.7 mU/L). Concentrations <18 pmol/L were considered suppressed; ≥18 pmol/L un-suppressed.Receiver operating characteristics (ROC) curves with determination of area under the curve (AUC) values were performed for the diagnostic performance of p-insulin in the diagnosis of CHI. Results: In the 61 samples from CHI patients, the median (range) p-insulin was un-suppressed in all diagnostic samples [90; 20-758 pmol/L (12.9; 2.9-109.1 mU/L)], while p-insulin was suppressed in all 15 samples from IKH patients [1.5; 1.5-9 pmol/L (0.21; 0.21-1.3 mU/L)]. The ROC AUC was 1.0 (95%CI. 1.0-1.0) for the diagnosis of CHI defined both by the clinic and by gold standard. The optimal p-insulin cut-off was 14.5 pmol/L (2.1 mU/L) or 12.5 pmol/L (1.8 mU/L), for CHI patients by use of a simultaneous p-glucose cut-off of <3.2 mmol/L (57.5 mg/dL; n=61), or 3.0 mmol/L (55 mg/dL; n=49), respectively. Conclusions: The sensitive insulin assay performed excellent in diagnosing CHI with a ROC AUC of 1.0. The use of a p-insulin cut-off of 13 pmol/L (1.86 mU/L) during a diagnostic hypoketotic hypoglycaemia test may establish the diagnosis of CHI without further diagnostic testing.

Control of ventricular rate in permanent atrial fibrillation: cardio-protec-tion and tissue hemodynamics

Objective To evaluate myocardial injury and tissue hemodynamics in elderly patients with permanent atrial fibrillation (AF) based on the achieved range of ventricular contraction rate (VCR).Materials and Methods This prospective, randomized, blind study included 75 patients aged ≥60 with permanent AF. All patients were prescribed bisoprolol as a VCR-reducing therapy. Patients were randomized to two groups according to the permuted-block design based on the range of resting VCR goal: group 1, 60-79 bpm (n=38) and group 2, 80-100 bpm (n=37). All patients also received perindopril and apixaban. Troponin I concentration was measured using the high-sensitivity assay (hsTn); parameters of tissue hemodynamics, including the mean blood flow velocity (Vm) and pulsatility index (PI), were measured using high-frequency ultrasound doppler flowmetry; echocardiographic indexes of left heart remodeling were recorded at baseline and after 6 month of VCR monitoring.Results Mean age of patients was 74±7 years. Medians [25th percentile; 75th percentile] of baseline hsTn concentrations were 10.2 [5.25; 21.2] ng / l in group 1 and 10.3 [5.4; 20.4] ng / ml in group 2 (p=0.91). 89.5 % of patients in group 1 and 100 % of patients in group 2 achieved the VCR range goal. At 6 month, resting VCRs were 70±4 bpm in group 1 (n=34) and 88±5 bpm in group 2 (n=37) (p1, p2&lt;0.001). According to echocardiographic data significant progression of myocardial remodeling was not observed. Concentrations of hsTn significantly decreased in both groups but the decrease was more pronounced in group 1, to 8.0 [4.13; 17.23; p1&lt;0.001] ng / l vs. 9.2 [4.8, 17.5] ng / l in group 2 (р1, p2&lt;0.001). A weak direct correlation was found between the VCR decrease and hsTn concentration (rs=0.44; p=0.009 in group 1, and rs=0.41; p=0.01 in group 2); regression coefficient was 0.78 at 95 % confidence interval (CI), from 0.21 to 1.3 (p=0.009) in group 1, and 0.14 at 95 % CI, from 0.04 to 0.24 (p=0.007) in group 2. Vm values were increased to 2.93±0.10 (p&lt;0.001) and 3.21±0.09 cm / sec (p&lt;0.001) and PIs were decreased to 1.42±0.03 conv. units (p&lt;0.01) and to 1.34±0.02 conv. units (p&lt;0.001) in groups 1 and 2, respectively.Conclusion The treatment aimed at VCR control in patients older than 60 with permanent AF was associated with a positive dynamics of myocardial injury (hsTn) and tissue hemodynamics indexes (Vm и РI). This indicates a possibility for using these indexes for further improvement of managing such patients.

A universal multiplex assay format for ultra-sensitive cytokine quantification in human serum and plasma

Abstract Quantifying prospective biomarkers of immune signaling in biological samples requires high assay sensitivity. The Simoa Planar Array provides greater sensitivity than ELISA, but has a finite assay menu. This study aimed to develop a universal multiplex ‘homebrew’ planar immunoassay format for development of high sensitivity assays for any analyte. Anchor antibodies specific for a peptide tag were microprinted in microwell plates. The peptide tags were conjugated to analyte-specific capture antibodies via maleimide chemistry, enabling high affinity binding of capture to anchor antibody. Bound sample antigen was sandwiched between the peptide-tagged capture and biotinylated detector antibodies; chemiluminescent detection was used to quantify signal. This ‘homebrew’ approach successfully detected sub picomolar levels of cytokines and chemokines in human serum and plasma in a multiplex assay measuring IL-5, IL-6, IL-10, IL-22, TNFa. When compared to commercially available planar assays the universal planar assay format provided comparable LLOQ and LOD with sample correlation (R2) &amp;gt;0.97 for all analytes tested. The universal planar assay was used to successfully screen multiple antibody pairs for human and mouse biomarkers and to develop robust assays using mouse, rabbit or goat monoclonal and polyclonal antibodies. This universal multiplex planar homebrew assay format combines the accuracy, reproducibility and high sensitivity of Simoa Planar Array technology with total analyte flexibility, and provides assay developers a simple and versatile method to rapidly develop robust high-sensitivity multiplex assays for any analyte of their choice.

Cross-platform comparison of highly sensitive immunoassay technologies for cytokine markers: Platform performance in post-traumatic stress disorder and Parkinson’s disease

There is mounting evidence of systemic inflammation in post-traumatic stress disorder (PTSD) and Parkinson’s disease (PD), yet inconsistency and a lack of replicability in findings of putative biological markers have delayed progress in this space. Variability in performance between platforms may contribute to the lack of consensus in the biomarker literature, as has been seen for a number of psychiatric disorders, including PTSD. Thus, there is a need for high-performance, scalable, and validated platforms for the discovery and development of biomarkers of inflammation for use in drug development and as clinical diagnostics. To identify the best platform for use in future biomarker discovery efforts, we conducted a comprehensive cross-platform and cross-assay evaluation across five leading platform technologies. This initial assessment focused on four cytokines that have been implicated PTSD – interleukin (IL)-1β, IL-6, tumor necrosis factor (TNF)-α, and interferon (IFN)-γ. To assess platform performance and understand likely measurements in individuals with brain disorders, serum and plasma samples were obtained from individuals with PTSD (n = 13) or Parkinson’s Disease (n = 14) as well as healthy controls (n = 5). We compared platform performance across a number of common analytic parameters, including assay precision, sensitivity, frequency of endogenous analyte detection (FEAD), correlation between platforms, and parallelism in measurement of cytokines using a serial dilution series. The single molecule array (Simoa™) ultra-sensitive platform (Quanterix), MESO V-Plex (Mesoscale Discovery), and Luminex xMAP® (Myriad) were conducted by their respective vendors, while Luminex® and Quantikine® high-sensitivity ELISA assays were evaluated by R&D System’s Biomarker Testing Services. The assay with the highest sensitivity in detecting endogenous analytes across all analytes and clinical populations (i.e. the highest FEAD), was the Simoa™ platform. In contrast, more variable performance was observed for MESO V-plex, R&D Luminex® and Quantikine®, while Myriad’s Luminex xMAP® exhibited low FEAD across all analytes and samples. Simoa™ also demonstrated high precision in detecting endogenous cytokines, as reflected in < 20 percent coefficient of variance (%CV) across replicate runs for samples from the healthy controls, PTSD patients, and PD patients. In contrast, MESO V-Plex, R&D Luminex® and Quantikine® had variable performance in terms of precision across cytokines. Myriad Luminex xMAP® could not be included in precision estimates because the vendor did not run samples in duplicate. For cross-platform performance comparisons, the highest cross-platform correlations were observed for IL-6 such that all platforms – except for Myriad’s Luminex xMAP® – had strong correlations with one another in measurements of IL-6 (r range = 0.59 – 0.86). For the other cytokines, there was low to no correlation across platforms, such that reported measurements of IL-1β, TNF-α, and IFN-γ varied across assays. Taken together, these findings provide novel evidence that the choice of immunoassay could greatly impact reported cytokine findings. The current study provides crucial information on the variability in performance between platforms and across immunoassays that may help inform the selection of assay in future research studies. Further, the results emphasize the need for performing comparative evaluations of immunoassays as new technologies emerge over time, particularly given the lack of reference standards for the quantitative assessments of cytokines.

The European Biological Variation Study (EuBIVAS): weekly biological variation of cardiac troponin I estimated by the use of two different high-sensitivity cardiac troponin I assays.

Background Cardiac troponins (cTn) are specific markers for cardiac damage and acute coronary syndromes. The availability of new high-sensitivity assays allows cTn detection in healthy people, thus permitting the estimation of biological variation (BV) of cTn. The knowledge of BV is important to define analytical performance specifications (APS) and reference change values (RCVs). The aim of this study was to estimate the within- and between-subject weekly BV (CVI, CVG) of cTnI applying two high-sensitivity cTnI assays, using European Biological Variation Study (EuBIVAS) specimens. Methods Thirty-eight men and 53 women underwent weekly fasting blood drawings for 10 consecutive weeks. Duplicate measurements were performed with Singulex Clarity (Singulex, USA) and Siemens Atellica (Siemens Healthineers, Germany). Results cTnI was measurable in 99.4% and 74.3% of the samples with Singulex and Atellica assays, respectively. Concentrations were significantly higher in men than in women with both methods. The CVI estimates with 95% confidence interval (CI) were for Singulex 16.6% (15.6-17.7) and for Atellica 13.8% (12.7-15.0), with the observed difference likely being caused by the different number of measurable samples. No significant CVI differences were observed between men and women. The CVG estimates for women were 40.3% and 36.3%, and for men 65.3% and 36.5% for Singulex and Atellica, respectively. The resulting APS and RCVs were similar for the two methods. Conclusions This is the first study able to estimate cTnI BV for such a large cohort of well-characterized healthy individuals deriving objective APS and RCV values for detecting significant variations in cTnI serial measurements, even within the 99th percentile.

A TaqMan-MGB real-time RT-PCR assay with an internal amplification control for rapid detection of Muscovy duck reovirus

A real-time reverse transcriptase-polymerase chain reaction (RT-PCR) for the detection of Muscovy duck reovirus (MDRV) RNA in clinical samples is described. The assay is based on TaqMan-MGB technology, consisting of two primers and one probe labeled with the reporter dye 6-carboxyfluorescein that binds selectively to the sigma B-protein gene of MDRV. This technique also includes an Internal Positive Control (IPC). The real-time RT-PCR assay was able to detect MDRVs, whereas other common waterfowl-origin viral pathogens were not recognised by the established oligonucleotide set, thus showing that the test was specific for MDRV. The sensitivity of the assay was 2.83 × 101 copies/μL and was 100 times higher than that of the conventional RT-PCR. The variation coefficients of intra-assay and inter-assay were less than 1.5% which verified sufficient repeatability of this assay.The use of β-actin mRNA as an IPC in order not to reduce the efficiency of the assay was adopted. The detection for 100 clinical samples showed that the positive rate of the established TaqMan-MGB real-time RT-PCR method was 87% (87/100), while the positive rate of the conventional RT-PCR was 83% (83/100), with the coincidence rate was 97.14%. Sensitivity and positive rate for clinical samples of TaqMan fluorescent quantitative RT-PCR were higher than conventional RT-PCR. The high specificity, sensitivity, and rapidity TaqMan-MGB real-time RT-PCR assay with the use of IPC to monitor for false negative results can make this method suitable for the pathogenic surveillance and epidemiological investigation of MDRV infection.

Improving individualized monitoring strategy for Rituximab treatment in patients with Neuromyelitis Optica Spectrum Disorders: development of a high sensitive ddPCR assay for CD19 mRNA quantification (4123)

Improving individualized monitoring strategy for Rituximab treatment in patients with Neuromyelitis Optica Spectrum Disorders: development of a high sensitive ddPCR assay for CD19 mRNA quantification (4123)

Comparative analysis of targeted next-generation sequencing panels for the detection of gene mutations in chronic lymphocytic leukemia: an ERIC multi-center study.

Next-generation sequencing (NGS) has transitioned from research to clinical routine, yet the comparability of different technologies for mutation profiling remains an open question. We performed a European multicenter (n=6) evaluation of three amplicon-based NGS assays targeting 11 genes recurrently mutated in chronic lymphocytic leukemia. Each assay was assessed by two centers using 48 pre-characterized chronic lymphocytic leukemia samples; libraries were sequenced on the Illumina MiSeq instrument and bioinformatics analyses were centralized. Across all centers the median percentage of target reads ≥100x ranged from 94.2-99.8%. In order to rule out assay-specific technical variability, we first assessed variant calling at the individual assay level i.e., pairwise analysis of variants detected amongst partner centers. After filtering for variants present in the paired normal sample and removal of PCR/sequencing artefacts, the panels achieved 96.2% (Multiplicom), 97.7% (TruSeq) and 90% (HaloPlex) concordance at a variant allele frequency (VAF) >0.5%. Reproducibility was assessed by looking at the inter-laboratory variation in detecting mutations and 107 of 115 (93% concordance) mutations were detected by all six centers, while the remaining eight variants (7%) were undetected by a single center. Notably, 6 of 8 of these variants concerned minor subclonal mutations (VAF <5%). We sought to investigate low-frequency mutations further by using a high-sensitivity assay containing unique molecular identifiers, which confirmed the presence of several minor subclonal mutations. Thus, while amplicon-based approaches can be adopted for somatic mutation detection with VAF >5%, after rigorous validation, the use of unique molecular identifiers may be necessary to reach a higher sensitivity and ensure consistent and accurate detection of low-frequency variants.

Epidermal growth factor receptor mutations in non-small cell lung cancer undetected by high-sensitivity allele-specific real-time polymerase chain reaction-based assays.

Identifying epidermal growth factor receptor (EGFR) mutation status is critical for planning lung cancer treatment. Sanger sequencing detects both known and novel mutations but shows poor sensitivity. High-sensitivity allele-specific real-time polymerase chain reaction (ASRP)-based assays offer quick and reliable results, but may overlook uncommon mutations. We aimed to define the rate at which high-sensitivity ASRP-based assays missed uncommon EGFR mutations. Non-small cell lung cancer specimens that were diagnosed as EGFR wild-type (EGFR-WT) by high-sensitivity ASRP-based assays and had residual DNA samples were sent for Sanger sequencing. Patient characteristics and clinical features were evaluated by chart review, and outcomes of EGFR-tyrosine kinase inhibitor (EGFR-TKI) therapy were studied. Hundred DNA specimens diagnosed by high-sensitivity ASRP-based assays as EGFR-WT were rechecked by Sanger sequencing. Two samples which were re-biopsy specimens from patients with EGFR mutations were excluded from the analysis. Sanger sequencing was failed in 24 samples. Among the remaining 74 samples, 6 (8.1%) had EGFR mutations-one exhibited exon 19 deletion (delT751_I759insS), two exhibited substitution mutations (S768I+V769L and L861Q), and three exhibited exon 20 insertions (N771_P772insN, P772_H773insHP, and H773_V774insAH). Only the patient with the exon 19 deletion had received EGFR-TKI therapy. Although the best tumor response was only stable disease, this was maintained for >10 months. High-sensitivity ASRP-based assays can overlook uncommon mutations. This detection failure rate is worth noting, especially when treating patients from regions known to have a high prevalence of EGFR mutation. Patients carrying uncommon mutations may still benefit from EGFR-TKI therapy.

Troponin elevation: is it ischemia?

Cardiac troponin is the preferred biomarker for myocardial infarction, thanks to its sensitivity and absolute specificity for the heart. The availability of high-sensitivity assays (hs-cTnT and hs-cTnI), capable of measuring with excellent analytical precision very low levels of circulating troponin, raised the issue of whether transient ischemia is a sufficient stimulus for troponin release. For this purpose, in a series of patients submitted to a stress test (exercise ECG/echo test; dipyridamole echo test; dobutamine echo test), we measured plasma levels of hs-cTnT at baseline and 6 hours after the end of the test. Plasma concentrations of hs-cTnT significantly increased in the vast majority of patients after the test. Significant elevations were documented in response to each stressor, regardless of the test result, after both positive and negative tests. Moreover, troponin significantly increased in response to the stress, both in patients with and in patients without obstructive coronary artery disease. Despite a good sensitivity (80% and 89%), troponin showed a very low specificity (32% and 47%) for stress-induced ischemia and coronary artery disease, respectively. Myocardial release of troponin is a multifactorial process, mediated not only by cardiomyocyte necrosis, but also through several different mechanisms such as myocardial ischemia, increase in cardiac work, and hemodynamic overload. Transient elevation of high sensitivity cardiac troponin is not a useful tool for detecting spontaneous or stress-induced ischemia. L

One-Step Digital Immunoassay for Rapid and Sensitive Detection of Cardiac Troponin I.

A rapid and sensitive method to detect cardiac troponin I (cTnI) in human blood is critical to the diagnosis and treatment of acute myocardial infarction (AMI). Here, we describe a simple one-step digital immunoassay for single-molecule detection without washing steps. A sample containing cTnI mixed with detection antibody-conjugated gold nanoparticles (AuNPs) is added to a capture antibody-coated sensor surface and the formation of the antibody-cTnI-antibody sandwich is detected by digitally counting the binding of the individual gold nanoparticles to the sensor surface in real time using a bright-field optical imaging setup together with a differential imaging algorithm. The digital immunoassay detects cTnI in undiluted human plasma, which achieves a detection limit of 5.7 ng/L within a detection time of only 10 min, which meets the requirement of current clinical high-sensitivity troponin assay (∼70 ng/L cutoff). We anticipate that the one-step and real-time digital immunoassay can be applied to the detection of other disease biomarkers in blood.

Diagnostic utility of point of care high sensitive troponin-i assay for early diagnosis of acute myocardial infarction in patients presenting with acute onset chest pain in emergency departments. The early heart study

Background: An early diagnosis of myocardial infarction is highly important in the Emergency Department (ED). It facilitates rapid decision making and treatment and therefore improves the outcome in patients presenting with symptoms of chest pain. Aims and objectives: To study diagnostic utility of new point of care high sensitive troponin-I assay in early diagnosis of acute myocardial infarction in patients presenting with acute chest pain.

From CANTOS to CIRT to COLCOT to Clinic: Will All Atherosclerosis Patients Soon Be Treated With Combination Lipid-Lowering and Inflammation-Inhibiting Agents?

From CANTOS to CIRT to COLCOT to Clinic: Will All Atherosclerosis Patients Soon Be Treated With Combination Lipid-Lowering and Inflammation-Inhibiting Agents?

Abstract P194: Proteomic Analysis of Cardiac Troponin I And T in Older Adults Without Cardiovascular Disease

Background: Minute elevations in cardiac troponins detected with high sensitivity assays are associated with cardiovascular disease (CVD) and mortality. The biology underlying these associations is uncertain. We identified plasma proteins associated with cardiac troponins and assessed the relation of troponin-associated proteins with CVD and all-cause mortality in adults without prevalent CVD. Methods: We included 3451 adults (62% female, 18% black, mean age: 75 years) in ARIC at Visit 5 (2011-2013). High sensitivity assays were used to measure troponins (hs-cTnI [Abbott], hs-cTnT [Roche]). We used linear regression to model the association of log2-transformed troponin and plasma proteins (SomaLogic aptamer assay). Upstream regulators of the troponin-associated proteins were identified using Ingenuity Pathway Analysis. Elastic net was used to extract troponin-associated protein signatures for principal component analysis and Cox models of CVD and all-cause mortality (median follow-up: 6 years). Results: Of the 4326 plasma proteins, there were 281 significantly (p&lt;1.16e-5) associated with hs-cTnI and 576 with hs-cTnT (adjusted for age and sex). In models adjusted for CVD risk factors, there were 99 proteins associated with hs-cTnI and 118 associated with hs-cTnT (Figure), with 65 in common; the top upstream regulator associated with both hs-cTnI (p = 0.001) and hs-cTnT (p=0.006) was myocardin (MYOCD), a transcriptional activator in smooth and cardiac muscles. The first principal component derived from the troponin-associated proteins was significantly associated with both all-cause and CVD mortality. In models with both troponins and the first principal component, hs-cTnI, but not hs-cTnT, remained significantly associated with both outcomes (Table). Conclusions: Hs-cTnI and hs-cTnT have overlapping, but distinct, proteomic signatures. The underlying proteomic signature explains most of the mortality risk associated with hs-cTnT, but not hs-cTnI.

Hot Off the Press: Troponin Testing and Coronary Syndrome in Geriatric Patients With Nonspecific Complaints: Are We Overtesting?

Elderly patients frequently present to the emergency department with nonspecific complaints and it falls to the emergency physicians to determine whether a workup for acute coronary syndrome is warranted. We discuss a study by Wang et al published in Academic Emergency Medicine, January 2020, looking at the value of troponin testing in elderly patients presenting to the emergency department with nonspecific complaints. We provide critical analysis of the article, and summarize the social media discussion and a podcast in which the authors discuss their work.

Biological variation of cardiac troponins in chronic kidney disease.

Background Patients with chronic kidney disease often have increased plasma cardiac troponin concentration in the absence of myocardial infarction. Incidence of myocardial infarction is high in this population, and diagnosis, particularly of non ST-segment elevation myocardial infarction (NSTEMI), is challenging. Knowledge of biological variation aids understanding of serial cardiac troponin measurements and could improve interpretation in clinical practice. The National Academy of Clinical Biochemistry (NACB) recommended the use of a 20% reference change value in patients with kidney failure. The aim of this study was to calculate the biological variation of cardiac troponin I and cardiac troponin T in patients with moderate chronic kidney disease (glomerular filtration rate [GFR] 30–59 mL/min/1.73 m2). Methods and results Plasma samples were obtained from 20 patients (median GFR 43.0 mL/min/1.73 m2) once a week for four consecutive weeks. Cardiac troponin I (Abbott ARCHITECT® i2000SR, median 4.3 ng/L, upper 99th percentile of reference population 26.2 ng/L) and cardiac troponin T (Roche Cobas® e601, median 11.8 ng/L, upper 99th percentile of reference population 14 ng/L) were measured in duplicate using high-sensitivity assays. After outlier removal and log transformation, 18 patients’ data were subject to ANOVA, and within-subject (CVI), between-subject (CVG) and analytical (CVA) variation calculated. Variation for cardiac troponin I was 15.0%, 105.6%, 8.3%, respectively, and for cardiac troponin T 7.4%, 78.4%, 3.1%, respectively. Reference change values for increasing and decreasing troponin concentrations were +60%/–38% for cardiac troponin I and +25%/–20% for cardiac troponin T. Conclusions The observed reference change value for cardiac troponin T is broadly compatible with the NACB recommendation, but for cardiac troponin I, larger changes are required to define significant change. The incorporation of separate RCVs for cardiac troponin I and cardiac troponin T, and separate RCVs for rising and falling concentrations of cardiac troponin, should be considered when developing guidance for interpretation of sequential cardiac troponin measurements.

A simple aptamer-based colorimetric assay for rapid detection of C-reactive protein using gold nanoparticles

C-reactive protein (CRP) is a clinical biomarker for inflammatory diseases. In this work, we present a simple and fast colorimetric method for CRP detection that employs citrate–capped gold nanoparticles (AuNPs) and a CRP-binding aptamer as sensing elements. The aptamer consisted in a guanine rich single-stranded DNA (ssDNA) that adsorbs onto the surface of the AuNPs. In the presence of the CRP, the ssDNA releases from the AuNPs surface to interact preferentially with the protein to form guanine-quadruplexes. The exposure of the unprotected AuNPs to buffer salts leads to aggregation and subsequent color change from red-wine to blue-purple that was readily seen by the naked eye. The AuNPs aggregation was monitored using UV–Vis spectroscopy and the CRP concentration in the samples could be correlated with the aggregation ratio (A670nm/A520nm). A linear sensing range of 0.889–20.7 μg/mL was found. The detection limit (LOD) was 1.2 μg/mL which is comparable to the typical clinical cutoff concentration in high-sensitivity CRP assays (1 μg/mL) and lower than the detection limit of nephelometric methods used in clinical practice. This method can provide a fast (5 min analysis time), simple, and sensitive way for CRP detection, with negligible interference of bovine serum albumin (BSA) up to concentrations of 100 nM.

High-sensitivity cardiac troponin assays for cardiovascular risk stratification in the general population.

Cardiac troponins (cTns) I and T have long been the most successful cardiac-specific circulating biomarkers in cardiovascular (CV) medicine, having changed dramatically the diagnosis of acute myocardial infarction, while being independent predictors of outcome in several cardiac conditions and non-cardiac conditions. The latest-generation high-sensitivity (hs) cTn assays demonstrate both enhanced diagnostic performance and improved analytical performance, with the ability to measure detectable concentrations in a substantial proportion of the asymptomatic and presumably healthy populations. Given this unique analytical feature, recent evidence suggests that hs-cTn can be used for the stratification of CV risk in the general population. High-sensitivity cTn predicts future CV events, are responsive to preventive pharmacological or lifestyle interventions, change in parallel to risk modifications, and offer incremental risk prediction when added to well-established prognosticators. The implementation of CV risk stratification and prevention strategies incorporating hs-cTn requires further investigation to define the optimal target populations, timing of measurement, and preventive interventions.

Early kinetic profiles of troponin I and T measured by high-sensitivity assays in patients with myocardial infarction

The early concentration kinetic profiles of cardiac troponin in patients with non-ST-elevated myocardial infarction (NSTEMI) measured by high-sensitivity cardiac troponin I (hs-cTnI) and T (hs-cTnT) assays have not been described.In intermediate-to-high-risk of NSTEMI patients we measured serial cTn concentrations on ED arrival, at 1, 2, 3, 6–12, 24 and 48-hours with hs-cTnI and hs-cTnT assays. Log-normal curves were fitted to concentrations from time from symptom onset, and the time to rule-out decision thresholds estimated (hs-cTnI: 2 ng/L and 5 ng/L; hs-cTnT: 5 ng/L).Among 164 patients there were 58 NSTEMI. The hs-cTnI to hs-cTnT ratio increased linearly over the first 6–12 h following symptom onset. The estimated times from symptom onset to the 2 ng/L and 5 ng/L thresholds for hs-cTnI were 1.8 (0.1–3.1) and 1.9 (1.1–3.5) hours, and to the 5 ng/L threshold for hs-cTnT 1.9 (1.1–3.8) hours. The estimated time to exceed 5 ng/L was ≥3 hours in 32.6% (95%CI: 20.0% to 48.1%) cases for hs-cTnI and 33.3% (19.6% to 50.0%) for hs-cTnT.cTnI concentrations increased at a much more rapid rate than cTnT concentrations in patients with NSTEMI. Concentrations of a high proportion of patients took longer than 3 hours from symptom onset to exceed the 5 ng/L rule-out decision threshold.