Clinical chemistry and laboratory medicine最新文献

{"title":"Response to the editorial by Karl Lackner.","authors":"Marith van Schrojenstein Lantman, Marc Thelen","doi":"10.1515/cclm-2025-0334","DOIUrl":"https://doi.org/10.1515/cclm-2025-0334","url":null,"abstract":"","PeriodicalId":10390,"journal":{"name":"Clinical chemistry and laboratory medicine","volume":" ","pages":""},"PeriodicalIF":3.7,"publicationDate":"2025-07-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144741358","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":"Accelerating time from result to clinical action: impact of an automated critical results reporting system.","authors":"Elisa M Aponte, Amos J Shemesh, Taylor Kalomeris, Bulent Oral, Stanley Diaz, Yaxin Li, He S Yang, Melissa M Cushing, Zhen Zhao","doi":"10.1515/cclm-2025-0711","DOIUrl":"https://doi.org/10.1515/cclm-2025-0711","url":null,"abstract":"","PeriodicalId":10390,"journal":{"name":"Clinical chemistry and laboratory medicine","volume":" ","pages":""},"PeriodicalIF":3.7,"publicationDate":"2025-07-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144728424","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":"Isotope dilution-liquid chromatography-tandem mass spectrometry-based candidate reference measurement procedures for the quantification of total and free phenytoin in human serum and plasma.","authors":"Tobias Schierscher, Linda Salzmann, Neeraj Singh, Manuel Seitz, Janik Wild, Carina Schäfer, Friederike Bauland, Andrea Geistanger, Lorenz Risch, Christian Geletneky, Christoph Seger, Judith Taibon","doi":"10.1515/cclm-2024-0858","DOIUrl":"10.1515/cclm-2024-0858","url":null,"abstract":"<p><strong>Objectives: </strong>An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure (RMP) was developed and validated to accurately measure serum and plasma concentrations of total and free phenytoin.</p><p><strong>Methods: </strong>Quantitative nuclear magnetic resonance spectroscopy (qNMR) was used to determine the absolute content of the reference material, ensuring its traceability to SI units. The separation of phenytoin from potential unknown interferences was achieved with reversed-phase chromatography, utilizing a C8 column. A protein precipitation protocol was established for preparation of total phenytoin samples, while free phenytoin samples were prepared by membrane separation utilizing a commercially available ultrafiltration device. Assay validation and determination of measurement uncertainties was performed according to the guidelines of the Clinical and Laboratory Standards Institute, the International Conference on Harmonization, and the Guide to the Expression of Uncertainty in Measurement.</p><p><strong>Results: </strong>These RMPs demonstrated high selectivity and specificity, with no evidence of matrix effects, allowing quantification of total and free phenytoin in ranges of 0.640-48.0 μg/mL and 0.0800-4.80 μg/mL, respectively. Intermediate precision was <3.8 %, and repeatability was 1.4-3.8 %, over all concentration levels, for both forms of phenytoin. For total phenytoin, relative mean bias ranged from -2.7-0.3 % in native serum and from 0.0-1.1 % in lithium heparin plasma. Relative mean biases for free phenytoin were 3.5-4.1 % for both native serum and ultrafiltrates. Measurement uncertainties for single measurements and target value assignment were 1.8-2.5 % and 0.9-1.7 %, respectively, for total phenytoin. For free phenytoin, these measurement uncertainties were 2.0-3.9 % and 0.9-1.4 % for single measurements and target value assignment, respectively.</p><p><strong>Conclusions: </strong>We present a novel LC-MS/MS-based RMP for phenytoin in human serum and plasma that provides a traceable and reliable platform for the standardization of routine assays and evaluation of clinically relevant samples.</p>","PeriodicalId":10390,"journal":{"name":"Clinical chemistry and laboratory medicine","volume":" ","pages":""},"PeriodicalIF":3.8,"publicationDate":"2025-06-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144339969","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":"An isotope dilution-liquid chromatography-tandem mass spectrometry based candidate reference measurement procedure for the simultaneous quantification of 25-hydroxyvitamin D3 and 25-hydroxyvitamin D2 in human serum and plasma.","authors":"Kerstin Kandler, Neeraj Singh, Friederike Bauland, Elie Fux, Andrea Geistanger, Christian Geletneky, Judith Taibon","doi":"10.1515/cclm-2024-1138","DOIUrl":"10.1515/cclm-2024-1138","url":null,"abstract":"<p><strong>Objectives: </strong>An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure (RMP) for the quantification of 25-hydroxyvitamin D2 (25OHD2) and 25-hydroxyvitamin D3 (25OHD3) in human serum and plasma is presented.</p><p><strong>Methods: </strong>Quantitative Nuclear Magnetic Resonance (qNMR) spectroscopic methodology has been utilized to assign absolute content (g/g) and International System of Units (SI)-traceability to the reference materials used as primary calibrators. This RMP was developed for the simultaneous quantification of 25OHD2 and 25OHD3 in human samples, utilizing supported liquid extraction (SLE) clean-up and a two-dimensional heart-cut ID-LC-MS/MS method to minimize matrix effects and prevent the co-elution of 3-Epi-25OHD3 and 3-Epi-25OHD2. The method underwent validation in accordance with current guidelines. Selectivity was assessed using spiked samples. To evaluate potential matrix effects, a post-column infusion experiment and a comparison of standard line slopes were performed. A 5-day validation study was conducted to determine precision, accuracy and trueness of the method. Measurement uncertainty for reference value assignment was evaluated in line with the Guide to the Expression of Uncertainty in Measurement (GUM). Equivalence to Joint Committee on Traceability in Laboratory Medicine (JCTLM) listed RMPs was demonstrated through the participation in the CDC Vitamin D Standardization-Certification Program (VDSCP) as well as the RELA scheme.</p><p><strong>Results: </strong>The RMP enabled the quantification of 25OHD2 and 25OHD3 within the range of 1.50 ng/mL-180 ng/mL (3.64-436 nmol/L for 25OHD2 and 3.74-449 nmol/L for 25OHD3), without interference from their respective epimer and no evidence of matrix effects. Intermediate precision was determined to be ≤4.0 % for 25OHD2 and ≤3.6 % for 25OHD3, while repeatability was ≤3.3 % for 25OHD2 and ≤2.9 % for 25OHD3 across all concentration levels. The relative mean bias for the secondary reference materials varied from -1.0 to 1.1 %, regardless of the analyte. For the spiked samples, the relative mean bias ranged from -4.2 to 1.0 % for 25OHD2 and from -3.9 to 0.9 % for 25OHD3, irrespective of all levels and matrices. Expanded measurement uncertainties (k=2) for target value assignment (n=6) were ≤3.9 % for 25OHD2 and ≤3.2 % for 25OHD3. Participation in the VDSCP and the RELA scheme showed a good agreement with results from the JCTLM listed RMPs and laboratories.</p><p><strong>Conclusions: </strong>The RMP enables the accurate, precise and consistent determination of 25OHD3 and 25OHD2. The robust performance of this method supports standardization of routine assays and guarantees traceability in the measurement of individual patient samples.</p>","PeriodicalId":10390,"journal":{"name":"Clinical chemistry and laboratory medicine","volume":" ","pages":""},"PeriodicalIF":3.8,"publicationDate":"2025-06-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144332556","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":"An isotope dilution-liquid chromatography-tandem mass spectrometry-based candidate reference measurement procedure for the quantification of 17β-estradiol in human serum and plasma.","authors":"Myriam Ott, Tobias Santner, Neeraj Singh, Friederike Bauland, Daniel Köppl, Alexander Gaudl, Andrea Geistanger, Uta Ceglarek, Manfred Rauh, Christian Geletneky, Judith Taibon","doi":"10.1515/cclm-2024-1255","DOIUrl":"https://doi.org/10.1515/cclm-2024-1255","url":null,"abstract":"<p><strong>Objectives: </strong>A new candidate isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based reference measurement procedure (RMP) has been developed for the accurate and precise quantification of 17β-estradiol (E2) in human serum and plasma covering a measurement range from 0.400 to 5,000 pg/mL (1.47-18,357 pmol/L). To address this broad range, two separate methods were created: a high sensitivity (HS) method for concentrations between 0.400 and 5.00 pg/mL (1.47-18.4 pmol/L) and a standard range (SR) method for concentrations between 5.00 and 5,000 pg/mL (18.4-18,357 pmol/L).</p><p><strong>Methods: </strong>As the primary reference material, E2 (CRM 6004-a) from the National Metrology Institute of Japan was used to ensure traceability to the international system (SI). A two-dimensional heart-cut LC approach was utilized for LC-MS/MS analysis, employing a supported liquid extraction sample preparation protocol for the SR method and a liquid-liquid extraction protocol followed by derivatization for the HS method. Assay validation was conducted following current guidelines. Selectivity and specificity were assessed using spiked serum samples, while potential matrix effects were evaluated through a post-column infusion experiment and comparison of standard line slopes. Precision, accuracy, and trueness were determined using an extensive 5-day protocol. Standard measurement uncertainty was evaluated according to the Guide to the Expression of Uncertainty in Measurement (GUM), with three individual sample preparations performed on at least two different days. Equivalence with higher-order RMPs was demonstrated through participation in the CDC Steroid Hormones Standardization (HoSt) program.</p><p><strong>Results: </strong>The RMP enabled the quantification of E2 within the range of 0.400-5,000 pg/mL (1.47-18,357 pmol/L), demonstrating no interference from structurally related compounds and no evidence of matrix effects. The relative mean bias of the SR method ranged from -2.4 to 1.9 % across all levels, including secondary reference materials and spiked samples, whereas the HS method exhibited a mean bias ranging from -3.0 to 2.9 %. Expanded measurement uncertainties (k=2) for target value assignment ranged from 1.7 to 4.4 % for the SR method and were found to be ≤6.1 % for the HS method. The method's transferability was demonstrated in a comparison study at a second laboratory. Additionally, the candidate RMP exhibited excellent correlation and equivalence to JCTLM-listed RMPs through the CDC HoSt program.</p><p><strong>Conclusions: </strong>In summary, the ID-LC-MS/MS-based RMP accurately quantifies E2. Its robust performance makes it suitable for standardizing routine assays and measuring individual patient samples, ensuring traceability.</p>","PeriodicalId":10390,"journal":{"name":"Clinical chemistry and laboratory medicine","volume":" ","pages":""},"PeriodicalIF":3.8,"publicationDate":"2025-05-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144157139","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":"Candidate reference measurement procedure based on isotope dilution-two dimensional-liquid chromatography-tandem mass spectrometry for the quantification of androstenedione in human serum and plasma.","authors":"Myriam Ott, Friederike Bauland, Vanessa Fischer, Daniel Köppl, Alexander Gaudl, Andrea Geistanger, Uta Ceglarek, Manfred Rauh, Judith Taibon","doi":"10.1515/cclm-2024-1135","DOIUrl":"https://doi.org/10.1515/cclm-2024-1135","url":null,"abstract":"<p><strong>Objectives: </strong>Androstenedione (ASD) is a biomarker used in the diagnosis of hyperandrogenism and adrenal hyperplasia. Therefore, accurate measurement of serum ASD is pivotal in clinical settings. We aimed to develop a novel highly selective reference measurement procedure (RMP) based on isotope dilution-two dimensional-liquid chromatography-tandem mass spectrometry (ID-2D-LC-MS/MS) for the quantification of ASD in human serum/plasma.</p><p><strong>Methods: </strong>To achieve high selectivity and sensitivity, a two-dimensional heart-cut LC approach for LC-MS/MS and a supported liquid extraction sample preparation protocol were employed. Matrix effects were evaluated through a post-column infusion experiment and comparison of standard line slopes. Precision and accuracy were determined via a multi-day validation experiment. Reproducibility was assessed by comparing results from two independent laboratories, and measurement uncertainty (MU) was evaluated in compliance with current guidelines.</p><p><strong>Results: </strong>Our novel RMP proved effective for measuring ASD concentrations ranging from 0.00800 ng/mL to 12.0 ng/mL (0.0279 nmol/L to 41.9 nmol/L) and demonstrated matrix-independence. The relative mean bias varied between 0.6 and 2.2 % across different matrices and concentration levels. Intermediate precision was observed to be between 1.2 and 1.9 %. The expanded measurement uncertainty for ASD target value assignment ranged from 1.7 to 2.5 %, irrespective of sample concentration. Equivalence to the JCTLM-listed RMP was evaluated through a method comparison study, revealing a high degree of agreement (r≥0.997). Additionally, by comparing results from two independent laboratories, the transferability and reproducibility of the RMP were confirmed.</p><p><strong>Conclusions: </strong>This isotope dilution two-dimensional LC-MS/MS method can be used for the evaluation and standardization of routine assays and for measuring individual patient samples, ensuring traceability.</p>","PeriodicalId":10390,"journal":{"name":"Clinical chemistry and laboratory medicine","volume":" ","pages":""},"PeriodicalIF":3.8,"publicationDate":"2025-05-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144149618","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":"The accuracy of presepsin in diagnosing neonatal late-onset sepsis in critically ill neonates: a prospective study.","authors":"Cinzia Auriti, Domenico Umberto De Rose, Chiara Maddaloni, Lucilla Ravà, Ludovica Martini, Eleonora Di Tommaso, Paola Bernaschi, Emanuel Paionni, Ottavia Porzio, Fiammetta Piersigilli, Marco Iannetta, Andrea Dotta, Maria Paola Ronchetti","doi":"10.1515/cclm-2025-0128","DOIUrl":"10.1515/cclm-2025-0128","url":null,"abstract":"<p><strong>Objectives: </strong>The diagnostic accuracy of presepsin (P-SEP) in the newborn is still under evaluation.</p><p><strong>Methods: </strong>In a multicenter study, we studied the accuracy of P-SEP as a diagnostic marker of late-onset sepsis (LOS) in critical newborns with underlying disorders, to define the most accurate cut-off to distinguish infected from uninfected patients.</p><p><strong>Results: </strong>Sixty-nine/351 newborns without infections at admission developed LOS. The median P-SEP value at T0 (admission) was 518.0 ng/L (IQR 313.0-789.0), without significant differences related to underlying diseases (p=0.52). In neonates who developed LOS, P-SEP increased at the onset of infection (T1) (median: 816.0 ng/L) and after 24-48 h (median: 901.0 ng/L) compared with their value at admission (median: 560.0 ng/L) (p<0.01 and p=0.03, respectively). The area under the ROC curve at T1 was 0.71 (95 % CI 0.65-0.78) when all cases of sepsis were included in the analysis and increased to 0.74 (95 % CI 0.66-0.81) considering only confirmed sepsis. Approximately two-thirds of patients were correctly classified, setting the cut-off at 713 ng/L, with a negative predictive value of 89.0 %.</p><p><strong>Conclusions: </strong>At a cut-off of 713 ng/L, P-SEP has good accuracy in diagnosing LOS in critically ill newborns. In uninfected newborns, the median value of P-SEP is not influenced by any underlying pathology.</p>","PeriodicalId":10390,"journal":{"name":"Clinical chemistry and laboratory medicine","volume":" ","pages":"1876-1887"},"PeriodicalIF":3.7,"publicationDate":"2025-04-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143962217","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":"An isotope dilution-liquid chromatography-tandem mass spectrometry-based candidate reference measurement procedure for the quantification of cortisone in human serum and plasma.","authors":"Myriam Ott, Neeraj Singh, Marie Kubicova, Friederike Bauland, Daniel Köppl, Alexander Gaudl, Andrea Geistanger, Uta Ceglarek, Manfred Rauh, Christian Geletneky, Judith Taibon","doi":"10.1515/cclm-2024-1478","DOIUrl":"https://doi.org/10.1515/cclm-2024-1478","url":null,"abstract":"<p><strong>Objectives: </strong>Cortisone is an inert precursor/metabolite of the potent steroid hormone cortisol. Measurement of serum cortisone levels and the cortisol-cortisone ratio can be useful for the diagnosis of dysfunction in the regulation of cortisol levels (i.e., severe and subtle apparent mineralocorticoid excess, low-renin primary aldosteronism). Therefore, an isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC MS/MS)-based candidate reference measurement procedure (RMP) to quantify cortisone in human serum/plasma was developed and validated.</p><p><strong>Methods: </strong>Quantitative nuclear magnetic resonance (qNMR) was utilized to assign absolute content (g/g) and SI-traceability to reference materials used as primary calibrators. A supported liquid extraction sample preparation protocol as well as a two-dimensional heart-cut LC approach for LC-MS/MS analysis were employed to mitigate matrix effects and prevent co-elution of interferences. Selectivity was determined by analyzing a matrix sample containing the analyte, the internal standard and six potential interferents. A post-column infusion experiment and a comparison of standard line slopes were performed to evaluate matrix effects. An extensive protocol over five days was applied to determine precision, accuracy and trueness. Measurement uncertainty (MU) was evaluated in compliance with current guidelines.</p><p><strong>Results: </strong>This RMP is suitable for analyzing cortisone within the 0.0800-120 ng/mL (0.222-333 nmol/L) range, demonstrating selectivity, sensitivity and matrix independence. Intermediate precision was ≤3.4 %, repeatability was ≤2.9 % across all concentration levels and relative mean bias ranged from -3.7 to 2.8 % across all tested matrices and concentrations. Expanded MU (k=2) for target value assignment (n=6) ranged from 2.1 to 5.5 %, irrespective of concentration or sample type.</p><p><strong>Conclusions: </strong>This RMP allows for accurate and reproducible determination of cortisone in human serum and plasma. Implementation of this method supports routine assay standardization and patient sample measurement with confirmed traceability.</p>","PeriodicalId":10390,"journal":{"name":"Clinical chemistry and laboratory medicine","volume":" ","pages":""},"PeriodicalIF":3.8,"publicationDate":"2025-04-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143968284","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":"Clostebol detection after transdermal and transmucosal contact. A systematic review.","authors":"Vincenzo Giannicola Menditto, Giulia Rossetti, Alessia Ferrarini, Angela Peghetti, Maria Domenica Camerlingo, Giovanni Pomponio","doi":"10.1515/cclm-2025-0331","DOIUrl":"10.1515/cclm-2025-0331","url":null,"abstract":"<p><strong>Introduction: </strong>To analyze the available evidence about the correlation between the presence of detectable amounts of clostebol metabolites in urine and the transdermal or transmucosal contact of clostebol.</p><p><strong>Content: </strong>A systematic review was performed. A systematic search was conducted in PubMed/MEDLINE, Scopus, Web of Science and the Cochrane library databases. Criteria for including studies were clinical studies reporting: (i) adult subjects; (ii) detection of urine clostebol metabolites derived from transdermal or transmucosal contact of clostebol.</p><p><strong>Summary: </strong>Seven papers pertinent to our questions were found: 3 case reports, one experimental study and 3 case reports with an experimental section for a total of 32 subjects. The median concentration of urine clostebol's metabolite 4-chloro-androst-4-en-3α-ol-17-one, M1 was 0.5 ng/mL (range 0.086-4.000 ng/mL; 25%-75 % IQ: 0.5-0.9 ng/mL) and 8.1 ng/mL (range 1.0-36.6 ng/mL; 25%-75 % IQ: 2.8-22.0 ng/mL), in subjects with indirect and direct exposure of clostebol, respectively (p=0.005).</p><p><strong>Outlook: </strong>We found consistent data that the detection of M1 in urine can be reconcilable with a transdermal or transmucosal contact of clostebol. In the cases of indirect exposure, the urine concentrations of M1 seem to be far lower than the concentrations found in case of direct exposure.</p>","PeriodicalId":10390,"journal":{"name":"Clinical chemistry and laboratory medicine","volume":" ","pages":"1472-1480"},"PeriodicalIF":3.7,"publicationDate":"2025-04-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143977145","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":"Isotope dilution-liquid chromatography-tandem mass spectrometry-based candidate reference measurement procedures for the quantification of 24<i>(R)</i>,25-dihydroxyvitamin D2 and 24<i>(R)</i>,25-dihydroxyvitamin D3 in human serum and plasma.","authors":"Kerstin Kandler, Michael Stadlmeier, Neeraj Singh, Friederike Bauland, Andrea Geistanger, Christian Geletneky, Judith Taibon","doi":"10.1515/cclm-2024-1139","DOIUrl":"https://doi.org/10.1515/cclm-2024-1139","url":null,"abstract":"<p><strong>Objectives: </strong>Isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC MS/MS)-based candidate reference measurement procedures (RMPs) for the quantification of 24,25(OH)₂D2 and 24,25(OH)₂D3 in human serum and plasma are presented.</p><p><strong>Methods: </strong>Quantitative nuclear magnetic resonance (qNMR) spectroscopic methodology was utilized to assign absolute content (g/g) and SI-traceability to reference materials used as primary calibrators. For liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis a two-dimensional heart cut LC approach, in combination with a supported liquid extraction protocol, was established to mitigate matrix effects and prevent co-elution of interferences. Selectivity was determined by spiking the internal standards and similar compounds, in human serum. A post-column infusion experiment and comparison of standard line slopes was performed to evaluate matrix effects. Precision and accuracy were assessed via a multi-day validation experiment, utilizing certified secondary reference materials from the National Institute of Standards and Technology (NIST). Measurement uncertainty (MU) was evaluated per the Guide to the Expression of Uncertainty in Measurement (GUM). To demonstrate equivalence with the JCTLM-listed RMP, certified secondary reference materials were utilized. Additionally, a method comparison study was conducted with the 24,25(OH)₂D3 method used by the CDC Vitamin D Reference Laboratory.</p><p><strong>Results: </strong>The RMP allowed quantification of 24,25(OH)2D2 and 24,25(OH)2D3 within the range of 0.150-18.0 ng/mL (0.350-42.0 nmol/L 24,25(OH)₂D2 and 0.360-43.2 nmol/L 24,25(OH)₂D3) without interference from structurally-related compounds and no evidence of matrix effects. Intermediate precision was ≤2.3 % for 24,25(OH)₂D2 and ≤2.9 % for 24,25(OH)₂D3; repeatability was ≤1.4 % for 24,25(OH)₂D2 and ≤2.1 % for 24,25(OH)₂D3, across all concentration levels. The relative mean bias was -4.5 to 2.9 % for 24,25(OH)₂D2, and -3.7 to 3.6 % for 24,25(OH)₂D3. Expanded MU for reference value assignment for 24,25(OH)₂D2 and 24,25(OH)₂D3 for reference value assignment was ≤2.5 %, regardless of concentration level and sample type. Passing-Bablok regression revealed strong agreement between the 24,25(OH)₂D3 results from the candidate RMPs and those provided by the CDC Vitamin D Reference Laboratory.</p><p><strong>Conclusions: </strong>These RMPs permit accurate and reproducible determination of 24,25(OH)₂D2 and 24,25(OH)₂D3. Implementation of these methods supports routine assay standardization and patient sample measurement with confirmed traceability.</p>","PeriodicalId":10390,"journal":{"name":"Clinical chemistry and laboratory medicine","volume":" ","pages":""},"PeriodicalIF":3.8,"publicationDate":"2025-03-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143556017","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}