Clinical chemistry最新文献

{"title":"B-229 Interlaboratory study using spike-ins and control samples to assess sample extraction and sequencing biases for metagenomics workflows","authors":"Jason Kralj, Stephanie Servetas, Samuel Forry, Monique Hunter, Scott Jackson","doi":"10.1093/clinchem/hvaf086.617","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf086.617","url":null,"abstract":"Background Sequencing bias remains a major obstacle to comparing metagenomic sample analyses and data reuse. Methodological variables impact the data, causing significant challenges to interpretating results from otherwise similar studies. However, controls in the form of spike-ins and common samples provide mechanisms for comparing between workflows to characterize biases. These allow methodological discrepancies to be resolved ahead of interlaboratory studies, and/or provide data to potentially reconcile differences from individual workflows. We initiated a small two-part interlaboratory study (ILS) to examine two questions: (a) does DNA extraction (5 methods) impact apparent sample composition; and (b) do DNA library/sequencing protocols (3 methods) have biases? Methods For ILS-a, participants were given 8 total samples (4x sample #1, 1x samples #2-#5) consisting of human stool spiked with a mixture of whole cells (200k/uL total of S. aureus, S. enterica, E. coli, L. monocytogenes, P. aeruginosa) and DNA internal controls (34k genome/uL ea. A. hydrophila & L. pneumophila). Participants extracted the DNA and returned the samples to NIST for sequencing. For ILS-b, participants were given 8 total samples consisting of DNA extracted from the same human stool samples in (a), spiked with DNA internal controls at ∼50k copy/uL/strain and spike-ins at ∼75k copy/ul/strain (see above). Labs generated DNA libraries, sequenced the samples, and returned the fastq files to NIST for processing. For (a) and (b), kraken2 was used to taxonomically classify the reads, reporting at the genus level. Relative abundance (reads / total reads) and Normalized abundance (reads / internal control reads) were used to examine the spike-ins and native taxa across the 5 samples. Results ILS-a (extraction) showed significant extraction bias between no change and 5-fold, with the spike-ins and native taxa mimicking similar trends in Gram +/- behavior. ILS-b (DNA) also showed significant bias vs. genome GC-content from different DNA library preparations (see Figure). These biases were reproducible between labs. Within-lab reproducibility of the 4 sample #1 replicates was 10-16% (a) and 9-18% (b), and the spike-in controls’ normalized abundances were consistent within lab across the 5 samples. This showed that the biases were sample composition-independent, and the biases were both reproducible and systematic. Conclusion Spike-ins and common-sample controls elucidate biases (and harmonization) between workflows, and indicate where data will likely have comparability challenges. The biases observed with the spike-ins were similar to the native taxa, such that a small number of well-characterized organisms helped account for biases across many native taxa. Hence, even small numbers of spike-ins provide a useful tool for assessing method bias, and indicate when more thorough method characterization may improve data intercomparability.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"72 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145203354","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A-116 Mixing it up: Salvaging frozen specimens","authors":"Ximena Wise, Nicholas Evans, Clovis Sarmiento, Hong-Kee Lee, Robert Benirschke","doi":"10.1093/clinchem/hvaf086.114","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf086.114","url":null,"abstract":"Background Proper specimen collection is necessary to minimize pre-analytical errors. Analyte stability after freezing is often provided with FDA approvals but is done on an aliquot instead of in the specimen collection container. The aim of this study is to examine the effect of freezing due to improper storage and/or transport on serum and plasma analytes from centrifuged and gel-separated samples. We investigate whether mixing thawed specimens can salvage the sample, preventing unnecessary patient re-draws and reducing laboratory costs. Methods Thirty fresh plasma and serum specimens that were not required for clinical care were randomly chosen. Baseline values for TSH, vitamin B12, vitamin D3, total PSA and Comprehensive Metabolic Panel (CMP) were collected. Specimens were frozen at -20? C for 24 hours, mimicking the winter conditions in outpatient pickup boxes. Specimens were then thawed at room temperature for 90-120 minutes and re-tested. Then, they were recapped and gently mixed for 10 minutes and re-tested a third time. CLIA, CAP and RCPA total allowable error (TAE) guidelines were utilized to assess whether or not a significant change occurred, in at least one sample, between the baseline and mixed values. Additionally, the data were analyzed using both male and female reference intervals to see if the clinical interpretations changed significantly within our laboratory (>10% of samples affected). Results Both serum and plasma specimens were affected significantly when assessed by either TAE and/or reference range standards. For the frozen, non-mixed plasma samples, the majority of the analytes (14/18, 78%)—except TPSA, Vitamin B12, TSH, and Alkaline Phosphatase—were outside TAE standards. However, upon mixing, eight of these analytes corrected themselves, leaving only six analytes (Potassium, Glucose, Total Bilirubin, Bicarbonate, AST and ALT) in the out-of-range category. Three analytes (AST, Bicarbonate, and Potassium, 18%) were altered when using the female reference intervals. Whereas when using the male reference intervals, Bicarbonate, Potassium, and TPSA had alterations in interpretation (17%). Regarding the serum samples, all of the analytes in the frozen, non-mixed specimens (18/18, 100%) were out of range per TAE standards. After mixing, only eight of the analytes (8/18, 44%) had fallen back within range. Regarding a change in clinical interpretation, five analytes (Bicarbonate, Chloride, Potassium, Sodium, and Vitamin D, 29%) were altered when using the female reference intervals, whereas when using the male values, a total of 6 analytes (aforementioned plus Creatinine, 33% of specimens) had changes in interpretation. Conclusion Real-world conditions may create pre-analytical errors that impact analyte results and must be considered. Importantly, this study showed that despite an analyte being stable per the FDA approval on an aliquot, this was not always the case in the specimen collection container. Plasma samples w","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"28 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145202931","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A-182 Analysis of Sample Recollections in an Outpatient Laboratory: A 5-Year Big Data Study","authors":"Mateus Alexandre, Rachel Petrola Jorge Bezerra","doi":"10.1093/clinchem/hvaf086.176","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf086.176","url":null,"abstract":"Background Laboratory testing requires continuous improvement to ensure accurate results. Standardizing processes and quality indicators is essential for the efficient management of laboratory services and analyses. The pre-analytical phase is particularly vulnerable to variables that may compromise sample integrity and lead to laboratory errors. Inadequate pre-analytical conditions can affect test results, leading to sample recollection, which delays diagnosis and causes patient discomfort. Additionally, recollection results in wasted supplies, time, and human resources. This study aims to identify the main causes of recollection and the most affected tests to improve protocols and enhance the quality of clinical analyses. Methods The present study is a descriptive, cross-sectional, and retrospective analysis of recollection cases in a Brazilian laboratory from 2020 to 2024, utilizing big data. The analysis focused on identifying the causes of recollection and the most affected tests among 23,016,266 tests performed during this period. Results This study critically analyzed the LIS database of a laboratory in Brazil, identifying 9,880 recollection cases over the 5 years. The proportion of recollections among all laboratory tests performed was 0.04% in 2020 (n=1618/3,646,518); 0.05% in 2021 (n = 2,160/4,515,699), 0.05% in 2022 (n=2,222/4,760,254), 0.04% in 2023 (n = 2,042/4,901,612), and 0.04% in 2024 (n = 1,838/5,192,183). Among the most frequent reasons for recollection, result confirmation accounted for 37.8% (3,737/9,880) of cases, followed by insufficient sample volume in 16.2% (1,605/9,880). Samples collected in the wrong tube were the third most common cause, representing 8.7% (868/9,880), while samples outside stability conditions accounted for 5.6% (557/9,880), and hemolysis for 4.3% (429/9880). When grouping the reasons for recollection, issues related to sample collection were the most prevalent 37,5% (3705/9880), followed by problems in sample storage 10,8% (1071/9880) and transportation 4,7% (467/9880). Regarding the most affected tests, recollections for zinc measurement accounted for 13.64% (1,348/9,880) of cases, followed by complete blood count (CBC) at 5.45% (538/9,880), copper at 5.02% (496/9,880), aluminum at 4.19% (414/9,880), and potassium at 2.97% (293/9,880). Grouping tests by section, mineral assays represented the highest proportion of recollections, 25.23% (2,493/9,880), followed by CBC in 5.45% (538/9,880), vitamin assays in 4.96% (490/9,880), and ion analysis at 4.74% (468/9,880). Also, coagulation tests accounted for 2.54% (251/9,880) of recollections. Conclusion This five-year analysis of sample recollections has highlighted key areas for improvement in the pre-analytical phase. The study reveals that factors such as result confirmation, insufficient sample volume, and improper sample collection are the primary causes. Tests related to zinc, complete blood count (CBC), and mineral assays were the most frequently aff","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"1 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145202932","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A-028 Comparison of the Cardiac Troponin I-T-C complex and cTnI and cTnT in Distinguishing Myocardial Infraction at Presentation","authors":"Yi Zhang","doi":"10.1093/clinchem/hvaf086.028","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf086.028","url":null,"abstract":"Background The potential relationship between cardiac troponin (cTn) fragment composition and the etiologies of myocardial injury indicate that characterization of cTn composition or targeting specific cTn fragments may provide diagnostic insights beyond total cTn levels and improve the diagnosis of myocardial infarction (MI). We analyzed the ternary cTnI–cTnT–TnC complexes (ITC), total cTnI and total cTnT in the circulation and compared the performance of the novel ITC complex assays with the widely used high-sensitivity (hs)-cTnI and hs-cTnT in differentiating MI patients at presentation. Methods Plasma samples were collected from 1210 patients presenting to the emergency department (ED) with symptoms suggestive of MI. The final diagnosis was adjudicated based on all available clinical records. Major cTn forms in the circulation were quantified using hs-cTnI, hs-cTnT, and ITC complex assays including long-cTnT ITC complex assay and hs-total ITC complex assay. We compared the performance of the ITC complex assays with hs-cTnI and hs-cTnT based on the 99th percentile upper reference limits (URLs), evaluated their area under the receiver operating characteristic curve (AUC), and assessed their ability to differentiate MI patients using established cut-off values. Results Among the study cohort, 138 (11.4%) patients were diagnosed with MI. The concentration of all cTn forms were significant higher in MI patients compared to non-MI patients. 86% patients had long-cTnT ITC complex concentration below the 99th percentile URLs, with the sensitivity 0.818 (95%CI: 0.746–0.874), specificity 0.943 (95%CI: 0.927–0.955), NPV 0.975 (95%CI: 0.964–0.983) and PPV 0.649 (95%CI: 0.575–0.716). Diagnostic accuracy, quantified by AUC values, was 0.959 (95% CI: 0.942–0.975) for the long-cTnT ITC complex, comparable to hs-cTnI (0.953, 95% CI: 0.934–0.971) and superior to both the hs-total ITC complex (0.889, 95% CI: 0.858–0.920, P < 0.001) and hs-cTnT (0.927, 95% CI: 0.904–0.948, P < 0.001). Using established cut-off values, the long-cTnT ITC complex assay classified 10% of patients as rule-in and 52% as rule-out, with a sensitivity of 0.993 (95% CI: 0.960–0.998), NPV of 0.998 (95% CI: 0.991–0.999), specificity of 0.972 (95% CI: 0.960–0.980), and PPV of 0.760 (95% CI: 0.678–0.826), which was comparable to hs-cTnI and better than hs-total ITC complex and hs-cTnT. In patients with a short chest pain onset time, the long-cTnT ITC complex assay demonstrated improved discrimination and enhanced diagnostic performance, as reflected by higher AUC values and more effective patient classification based on the 99th percentile URLs and established cut-off values. Conclusion The long-cTnT ITC complex assay demonstrated performance comparable to hs-cTnI and superior to both the hs-total ITC complex and hs-cTnT in distinguishing MI at presentation. Notably, its diagnostic advantages were more pronounced in patients at early presentation after symptom onset, highlightin","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"18 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145202999","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"B-048 New technology for Erythrocyte Sedimentation Rate (ESR). Comparison between automated measure and modified Westergren method in pediatric population. Is a change of methodology possible?","authors":"Maria Inés Marcone, Macarena Ottobre Saborido, Rodrigo Arrieta, Ramiro Gosis, Nadia Kauzlauskas, Mariana Campal, Silvia Salvatierra","doi":"10.1093/clinchem/hvaf086.446","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf086.446","url":null,"abstract":"Background ESR is based on the principle that the sedimentation of red blood cells in plasma provides an indirect measure of the level of acute-phase proteins and therefore of inflammation, assessing the acute phase response in pathological conditions. Although the International Council for Standardization in Hematology (ICSH) considers the Westergren as the reference method, the larger sample volume required limits its use in pediatrics. The technical innovations significantly improved on the existing procedures. One of the main advantages of automated systems includes the use of ethylenediaminetetraacetic acid (EDTA) that increases sample stability and allows using a single sample for both ESR and other hematological tests, thus enhancing patient safety, optimizing the workflow and increasing personnel safety by using closed systems and shortening turnaround time. Due to the wide variety of methods available, all new technologies must be evaluated against the Westergren reference method before being introduced into clinical use. The aim of this study was to evaluate ESR values obtained by an automated analyzer which integrates measurement of ESR with routine complete blood count (CBC) compared with modified Westergren method (ESRw) currently used. Methods Outpatients and hospitalized pediatric patients (1-18 years) were included (n=120). The manual measurement (3.8% sodium citrate tubes, 1 ml, Vitis®) using the ESRw was performed according to the ICSH’s recommendations. The Mindray? BC 6800 Plus ESR was measured in 1.3 ml EDTAK3 tubes (Tecnon®), simultaneously with CBC. Samples were stored at room temperature and analyzed by a dedicated technologist within 4 hours of collection. The analyzer measures ESR value using near-infrared photometry to determine the degree of red blood cell aggregation that occurs in the first phase (rouleaux formation) of sedimentation. Reportable ESR parameter (ESRm) and the Research-Use-Only ESR-correction parameter (ESRmc, independent of hematocrit value) were performed. The data was statistically analyzed and compared among different methods: Pearson correlation, Bland-Altman and Passing-Bablok (p-value < 0.05 was considered statistically significant). Results Both ESRm and ESRmc methods show very good correlation with the reference method: 0.9081 and 0.9169 respectively. Although ESRm has a very good correlation, it shows an associated systematic and proportional error, which is not observed in ESRmc (Figure 1-4). Conclusion Automation of ESR measurement is currently an attractive alternative even more in the pediatric population, where the sample volume is usually a limitation and the turnaround time of the results is critical. Taking this into account, the new technology allows the same sample used for CBC and provides results in 90 seconds. However, validation and comparison against the Westergren reference method is required. If the change of methodology is established, the existence of a bias should be","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"10 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145203115","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A-094 Mindray G6PDH –An Efficient and Accurate Method for Identifying G6PD Deficiency Population","authors":"Jian Dai, Qinlan Liu, Kaixiong Xu, Jin Li, Anping Xu, Feng Wang","doi":"10.1093/clinchem/hvaf086.093","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf086.093","url":null,"abstract":"Background Glucose-6-phosphate Dehydrogenase (G6PD) testing is used for diagnosis of G6PD deficiency. World Health Organization (WHO) advises that an individual’s G6PD status should be determined to guide dose and duration of anti-malarial drugs, and there are still unmet needs for G6PD testing to early screen neonates. The existing G6PD screening methods usually require multiple steps of manual operation. The results of these methods are often impacted by the variability or errors in manual operation. Therefore, they are difficult to accurately identify the population with G6PD mutation. This study evaluated the performance of a novel fully-automated G6PD/Hb detection method (G6PDH), and compared its diagnostic value with other methods, so as to provide a more precise technical means for the early screening and diagnosis of the disease. Methods In this study, the automatic lysis mode of Mindray BS-2800M analyzer was used to realize the G6PD/Hb twin-tests in parallel, in which G6PD and Hb (hemoglobin) were detected simultaneously using the same blood sample, and the G6PD/Hb ratio was calculated, defined as G6PDH. The age and gender specific reference intervals with this method were established. Based on the results of Multiplex Melting Curve Analysis (MMCA, a genetic mutation detecting method), the diagnostic efficacy and accordance rate of G6PDH and other biochemical methods such as G6PD/6PGD, G6PD single enzyme method were analyzed. Results According to the guidance of WHO, the reference intervals of G6PDH in non-neonates were established as <2.31, =2.31, 2.31∼6.17, =6.17 for deficient males/females, normal males, intermediate females, and normal females. For neonates, they were <4.14, =4.14, 4.14∼11.03, =11.03, respectively. Compared with the results of MMCA method on 185 samples, the sensitivity and accuracy of the G6PDH method were 93.5% and 91.7% respectively, which were significantly higher than those of G6PD/6PGD method (74.0%, 74.4%) and G6PD single enzyme method (68.3%, 67.8%). The Youden index of G6PDH method was 0.81, higher than that of 0.74 and 0.63 by other two methods, respectively. And the missed diagnosis rate was 6.5%, lower than that of the other two methods (26.0%, 31.7%). Moreover, in 64 female patients with heterozygous mutation, the diagnose accordance rate of G6PDH was 87.5%, while the other two was 51.6% and 39.1%. Conclusion Mindray provides an innovative automated G6PDH method on BS-2800M to accurately identify G6PD deficiency population, and significantly reduces the missed diagnosis rate of high-risk population with G6PD deficiency, especially for female heterozygous mutation. The G6PDH method could be well used for the screening, diagnosis and monitoring of G6PD deficiency, and could be considered as the preferred method in clinical laboratory.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"75 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145203118","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A-121 Analytical performance evaluation of the next generation Abbott enzyme assays on the Alinity c system","authors":"Meshach Asare-Werehene, Pow Lee Cheng, Xiao Yan Wang, Marvin Berman, Vathany Kulasingam","doi":"10.1093/clinchem/hvaf086.117","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf086.117","url":null,"abstract":"Background Liver enzyme assays, including alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma-glutamyltransferase (GGT), and alkaline phosphatase (ALP), are essential in clinical laboratories for assessing liver function, diagnosing liver diseases, and monitoring treatment efficacy. Thus, accurate measurement of these enzyme activities is crucial for clinical diagnostics. Recently, Abbott introduced a next generation of Alinity and Architect enzyme assays, incorporating a multianalyte calibrator (Consolidated Chemistry Calibrator, ConCC) with extended stability. For these new formulation assays, both ConCC and calibration factors are offered for all the assays except GGT, which had better performance with the ConCC calibrator. These assays achieve high Sigma metrics on the Alinity c system, minimizing variability and errors. This study evaluated the analytical performance of these newly developed next generation assays on the Abbott Alinity c platform. Methods Using 2 levels of quality control (QC) material (Bio-Rad Chemistry UA) and 3 pooled patient samples, we assessed imprecision by measuring these 5 samples twice per day (morning and afternoon), for five days. Acceptable imprecision and bias were determined based on the Accreditation Canada Diagnostics (ACDx) recommendations. Linearity testing consisted of 6 levels of commercially available linearity materials, with 3 replicates per level. Method comparison between the next generation assays and on-market conventional assays were evaluated in duplicates using patient specimens (n= 145 – 155) on the Abbott Alinity c platform. Passing-Bablok and Bland-Altman plots were used for the method comparison analyses. The precision and bias (external standard) studies were used to calculate the Sigma-metric using the formula, Sigma-metric=(%TEa-|%bias|)/%CV. TEa was defined based on ACDx and Clinical Laboratory Improvement Amendments (CLIA) recommendations. Results The Alinity next generation assays demonstrated acceptable imprecision, meeting the ACDx goals of ±2 U/L for concentrations =40 U/L and ±4% for concentrations >40 U/L for AST, ALT and GGT, and for ALP at ±4 U/L for concentrations =100 U/L and ±4% for concentrations >100 U/L. Furthermore, these assays exhibited linearity across the six concentration levels tested. All next generation Alinity clinical chemistry enzyme assays showed a Pearson*s R value of 1.0, indicating a strong linear correlation. The linearity slope ranged from 0.76 (ALP2) to 1.04 (AST2) whereas the y-intercept ranged from –59.40 (GGT2) to 8.78 (AST2). The majority of next generation assays performed at or above 6 Sigma. Good agreements were observed between the on-market assay and the next generation (ConCC and factor calibrated) assays. Conclusion The Alinity next generation assays (ALT2, AST2, GGT2 and ALP2) demonstrated acceptable performance for precision and linearity, with good agreement with the conventional factor-based assays on the","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"1 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145203119","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A-238 Influence of different apolipoprotein B-depleted serum preparation methods on high-density lipoprotein cholesterol efflux capacity assays","authors":"Saki Izumi, Tsunehiro Miyakoshi, Rina Kawaguchi, Akira Yoshimoto, Minoru Tozuka, Ryunosuke Ohkawa","doi":"10.1093/clinchem/hvaf086.231","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf086.231","url":null,"abstract":"Background Cholesterol efflux capacity (CEC), a critical anti-atherosclerotic function of high-density lipoprotein (HDL), is a promising biomarker for cardiovascular disease (CVD) risk. However, conventional CEC assays utilizing cultured cells are complex, hindering clinical application. To address this limitation, we previously developed a cell-free CEC assay, the immobilized liposome-bound gel beads (ILG) method. Both the ILG and conventional methods employ apolipoprotein B (apoB)-depleted serum (BDS) as the measurement sample. However, the optimal BDS preparation method remains unclear, as various methods exist, including polyethylene glycol (PEG), heparin sodium (Hep), and dextran sulfate (Dex). This study aimed to investigate the impact of different BDS preparation methods on both ILG and conventional cell-dependent assays. Methods Three types of BDS were prepared by mixing serum (set as 100 parts by volume) with the following reagents: (1) 20% PEG 6000 (200 mM glycine solution, pH 7.4) at a ratio of 100:40, (2) 1.3 mg/mL Hep in saline and 1M MnCl2 at 100:40:50, and (3) 5% Dex solution and 2M MgCl2 at 100:2:2.5. After centrifugation, the supernatants were collected as PEG-BDS, Hep-BDS, and Dex-BDS, respectively. The apoB-containing precipitates were dissolved in saline. Total cholesterol (TC) and HDL-C concentrations of each supernatant and precipitate solution were determined enzymatically. Apolipoprotein profiles were analyzed using SDS-PAGE and Native-PAGE followed by western blotting. CEC was measured using both the ILG method and conventional assay employing THP-1 cells and BODIPY-labeled cholesterol. To assess ATP-binding cassette transporter A1 (ABCA1)-mediated CEC, cells were cultured with and without a Liver X receptor agonist to modulate ABCA1 expression. Results While the three BDS exhibited different profiles, these differences did not significantly affect CEC measurements using the ILG method. TC concentrations in PEG-BDS and Dex-BDS were comparable to HDL-C concentrations in the original serum, whereas Hep-BDS exhibited significantly higher TC levels. SDS-PAGE revealed no differences in apoA-I and apoE profiles across the BDS samples, and barely detected apoB in any BDS, with the detected amount corresponding to less than 10% of serum. ApoA-I was present in all precipitates, with no significant differences in its relative abundance. However, Native-PAGE demonstrated a distinct distribution of HDL particles in Hep-BDS compared to the original serum. Importantly, the ILG method yielded comparable CEC values between the original serum and all BDS. In contrast, the cell-dependent assay revealed a significant reduction in ABCA1-mediated CEC in PEG-BDS compared to the original serum (approximately 56%). Furthermore, the Hep-BDS interfered with the cell-dependent assay by causing turbidity in the medium. Conclusion This study demonstrates that the choice of BDS preparation method does not significantly influence CEC measurements using","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"94 7 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145203321","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A-107 Evaluating the Use of Albumin-Corrected Calcium Measurements in an Acute Care Hospital Network","authors":"Annie Ren, Purvi Patel, Linda Stevenson, Lisa Brown, Veronica Roberts, Nicholas Pellegrino, Saranya Arnoldo","doi":"10.1093/clinchem/hvaf086.105","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf086.105","url":null,"abstract":"Background Using albumin-corrected calcium remains a conventional practice for determining calcium homeostasis, particularly in patients with hypoalbuminemia. However, studies have demonstrated that relying on albumin-corrected calcium in patients with hypoalbuminemia can overestimate serum calcium levels. We evaluated the correlation between total calcium measurements (with or without albumin correction) and ionized calcium levels in patients with normal and low albumin levels. Our findings were valuable for providing physician education on discontinuing the report of albumin-corrected calcium in our health system. This change aims to improve patient care and optimize laboratory resources. Methods We analyzed six months of patient data for total calcium, with and without albumin correction, obtained across three hospitals (N=56465). The Payne formula [Corrected calcium (mmol/L) = total calcium (mmol/L) + 0.02 [40 - albumin (g/L)] was used to derive the albumin-corrected calcium. We assessed paired ionized calcium specimens, collected within 24 hours apart (N=3123). The linear relationship and clinical correlation were assessed between total calcium, with or without albumin correction, with ionized calcium levels. Results Linear regression showed comparably moderate correlation between uncorrected (R² = 0.75, slope= 1.58) and corrected total calcium (R² = 0.74, slope= 1.55) with ionized calcium levels. In patients with hypoalbuminemia (N=2609), the clinical concordance with ionized calcium was stronger with uncorrected calcium compared to albumin-corrected calcium (72% vs 53%). Moreover, the clinical concordance with low ionized calcium was significantly higher using uncorrected calcium, compared to corrected calcium (88% vs 20%). When the ionized calcium is normal, the concordance was similar between uncorrected and corrected calcium (62% vs 60%). Conclusion Our findings support the literature that relying on the interpretation of albumin-corrected calcium can overestimate calcium status. There is a high risk of missing true hypocalcemia, where the corrected calcium level is in the normal range while the ionized calcium level is low. The uncorrected calcium measurement was more reliable for determining calcium status and agrees better with ionized calcium, in the presence of hypoalbuminemia.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"39 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145202928","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"B-258 Development and Fit-for-Purpose Analytical Validation of a LC-MS/MS Method for the Quantification of pS129 Alpha-synuclein in Human Serum","authors":"Sarika Chauhan, Shamili Sammohi, Erpan Ahat, Gary Heady, Andrew Schade, Michael Hodsdon","doi":"10.1093/clinchem/hvaf086.645","DOIUrl":"https://doi.org/10.1093/clinchem/hvaf086.645","url":null,"abstract":"Background Parkinson’s Disease (PD) poses significant challenges due to its long progression period and overlapping pathology with other neurodegenerative diseases. The phosphorylation of alpha-synuclein at serine 129 (pS129) has been shown to play a crucial role in PD progression, and potentially serve as a diagnostic biomarker for PD. Methods This assay development and validation was conducted in the Clinical Diagnostics Laboratory department of Eli Lilly & Company using an Orbitrap Mass Spectrometer. The assay method uses a pS129-specific antibody for immunoprecipitation (IP), followed by sample clean-up with solid-phase extraction (SPE) and protein digestion using trypsin (Waters, RapiZyme). After digestion, tryptic peptides were separated, and the alpha-synuclein (Asyn) peptide (aa81-96) was quantified with a Parallel Reaction Monitoring (PRM) method. A five-point calibration curve (30, 100, 300, 1000, 2000 pg/mL) was prepared by spiking in pS129 Asyn protein in 2% SigMatrix (Sigma-Aldrich). Low and high concentration quality control (QC) samples were run alongside the validation samples on each plate. A heavy-labeled tryptic internal standard peptide was added to each sample for normalization. Data analysis was conducted using Skyline Software (MacCoss lab, University of Washington). Assay precision, accuracy, sensitivity, parallelism, dilution linearity, interference, and sample stability were evaluated in this analytical validation. Results Precision across 65 measurements from five unique samples (3 healthy and 2 diseased) ranging from 200 pg/mL to 1500 pg/mL showed total CV ranging between 13% to 16%, for individual samples. Spike recovery (Accuracy) was evaluated by adding recombinant pS129 Asyn protein to both healthy and diseased samples at concentrations of 0 pg/mL (No spike), 200 pg/mL, and 500 pg/mL. The percentage recovery (%RE) range for accuracy was between 106% to 132%. No interference was observed from non-phosphorylated Asyn protein, hemoglobin, or intralipid. The limit of blank was 47 pg/mL. Functional analytical sensitivity (lower limit of quantitation) was at 81 pg/mL. The stability study conducted over a period of seven days at 4°C showed %RE ranging from 97% to 122%. Samples remained stable for up to five freeze/thaw cycles at -20°C and -80°C. The final analytical measurement range was 80-2000 pg/mL. The assay showed good parallelism (up to 1:4 dilution, %RE: 107%-112%) and dilutional linearity (up to 1:4 dilution, %RE: 73%-103%). Conclusion Analytical validation shows that the assay method is robust for detecting pS129 Asyn in human serum, which can be crucial for research and clinical applications related to neurodegenerative diseases.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":"9 1","pages":""},"PeriodicalIF":9.3,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145203120","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}