Apolipoprotein standardization in multiple calibration laboratories requires equivalent results to value assign matrix-based reference and external quality assurance materials. A multiplexed LC-MS/MS-based reference measurement procedure (RMP) has been developed for serum apolipoproteins apo(a), apoA-I, apoB, apoC-I, apoC-II, apoC-III, and apoE. This study evaluates the transferability of the RMP between 3 calibration labs and determines the between-laboratory imprecision. Six periodic ring trial surveys were held. The study protocol, calibrators, internal standards, quality controls (QCs), and clinical samples (CSs) were shared among the laboratories. Intra-laboratory imprecision and inter-peptide comparisons evaluated intra-laboratory performance, while inter-laboratory imprecision evaluated equivalence between the calibration labs. Precision of the common bilevel QC monitored the level of harmonization over time. Intra-laboratory imprecision fulfilled predefined analytical performance, defined as repeatability <50% of the maximum allowable uncertainty (MAU) at minimal criteria. Median interlaboratory variation (CVbl) was 3.71%, 3.33%, 7.38%, 6.74%, 3.88%, and 3.90% for apoA-I, apoB, apoC-I, apoC-II, apoC-III, and apoE, respectively. For apo(a), CVbl was concentration (x) dependent following 206.32×x-0.899%. In QC samples, the average imprecision for all apolipoproteins decreased from 6.0% and 18.1% for QC1 and QC2, to 5.2% and 9.5%, indicating improvement of analytical performance of the network over time. This study shows the feasibility of transferring the multiplex apo LC-MS/MS-based RMP between laboratories. Predefined performance specifications were fulfilled for all seven apolipoproteins. Ongoing round-robin studies will ensure stable performance of the calibration labs required to accomplish an accurate value-base for apolipoprotein certification of commercial reagents.

Automated phlebotomy has the potential to improve patient outcomes and address phlebotomist workforce challenges. The Autonomous Blood Drawing Optimization and Performance Testing trial (ADOPT, ClinicalTrials.gov NCT05878483) assesses performance, safety, and patient experience of an autonomous robotic phlebotomy device (ARPD) that utilizes multimodal imaging. Patients were recruited from outpatient phlebotomy departments in the Netherlands. In cohort 1, analytical equivalence was assessed by comparing within-subject test results from ARPD-collected and manually collected samples using Passing-Bablok regression analysis.In cohort 2, the ARPD was assessed in routine use. The primary performance endpoint was first-stick success rate; the primary safety endpoint was the rate of adverse events. Exploratory endpoints assessed pain and patient preference. One hundred fifty-three patients were enrolled in cohort 1. No statistically significant differences were observed in activated partial thromboplastin time, prothrombin time, lactate dehydrogenase, aspartate transaminase, or platelet count, indicating analytical equivalence.A total of 1633 patients were enrolled in cohort 2. The overall first-stick success rate, in patients where the ARPD could identify a suitable vein, was 94.5% (95% CI, 93.3-95.5) and remained high in patients with difficult venous access (92.7%), obesity (97.4%), and age ≥65 years (93.4%). Adverse events, all mild, occurred at a rate of 0.6%. Ninety percent of patients reported far less (19%), less (32%), or similar pain (39%) compared with manual phlebotomy. Eighty-two percent indicated they would strongly prefer (16%) or prefer (31%) use of the ARPD in the future or had no preference (35%). Automated phlebotomy using the ARPD shows a favorable performance and safety profile and may improve patient experience. ClinicalTrials.gov Registration Number: NCT05878483.

Bisulfite conversion remains the gold standard method for DNA methylation analysis but has known limitations. Enzymatic conversion has recently been developed and overcomes some of these pitfalls. This study compared both methods by measuring known colorectal cancer (CRC) methylated biomarkers in colorectal tissues. Colorectal tumors and matched adjacent normal tissues were collected before treatment at surgery from 24 patients with stage I-IV CRC (n = 6 per stage), aged 46-81 years (50% male). Tissue DNA was extracted and converted using bisulfite and enzymatic conversion. Methylation levels of BCAT1, IKZF1, and SEPTIN9 were quantified by methylation specific quantitative PCR (MS-qPCR) as a percentage of the reference gene (ACTB). Biomarker positivity in tissues was tested using a series of percentage methylation thresholds between ≥1% and ≥20%, and percentile-based cutoffs ranging from 50th to 95th percentiles. The Kruskal-Wallis test compared the methylation levels between the conversion methods. There were no significant differences in methylation levels between the bisulfite and enzymatic conversion methods for normal tissue DNA (P > 0.05). In tumor tissues, IKZF1 and SEPTIN9 had significantly higher DNA methylation levels with bisulfite conversion compared to enzymatic conversion (P < 0.05), while BCAT1 showed no significant difference (P > 0.05). Across all thresholds, there were no significant differences in colorectal tissue positivity between conversion techniques (P > 0.05), except IKZF1 at the 50th percentile in normal tissues was significantly hypermethylated in enzymatically converted DNA (P < 0.05). Bisulfite conversion has higher DNA methylation levels for certain CRC biomarkers in tumor tissues, suggesting over-estimation of methylation that could affect biomarker specificity.

The clinical use and interpretation of cardiac troponins (cTn) in pediatric patients is heterogeneous. This review is based on an appraisal and synthesis of the available literature and discussions between International Federation of Clinical Chemistry and Laboratory Medicine Committee on Clinical Application of Cardiac Biomarkers members and pediatricians, cardiologists, emergency department physicians, and laboratorians. It aims to summarize current knowledge and best practices related to cTn measurements in pediatric patients, including which percentile should be applicable as the upper reference limit, and the influence of age, sex, and physiology on circulating concentrations. Preanalytical and analytical precautions, and a consensus on optimal interpretation of the reported results are discussed. Current studies report high cTn concentrations in healthy infants and newborns. The nadir is reached at 1-3 years at which time concentrations gradually increase, reaching adult levels through puberty. The incidence of myocardial injury is reported to be low and dependent on whether the 97.5th or 99th percentile is chosen as the upper reference limit. The main clinical indication for testing appears to be suspicion of myo/pericarditis or traumatic chest injury. On occasion, cTn is measured in cases of possible heart failure and/or myocardial ischemia. There is heterogeneity among the limited clinical studies published. Most of the data are retrospective, and the current suggestions for clinical practice are thus largely based on expert opinion. More research is needed regarding all aspects of cTn testing in children, and suggestions for future research are included at the end of this review.