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Good estimates of analytical, within-subject, and between-subject variation, from appropriately designed biological variability studies, allow the potential for a biomarker to diagnose and monitor disease to be assessed in addition to setting goals for test performance. Sample sizes for these studies require consideration of the numbers of participants (n1), observations per participant (n2), and replicates of each observation (n3). Little guidance exists to compute these values. Data was simulated to examine the effect of changes in each component of sample size on analytical, within-subject, and between-subject variability, and on common measures of variability including the coefficient of variation (CV), reference change values (RCV), and index of individuality (II). Simulations showed the precision of estimated results (CVs, RCVs, and IIs) for varying sample sizes. Greater numbers of participants increased the precision of estimated analytical, within-subject, and between-subject variability; increasing the number of observations per participant increases the precision of estimates of analytical and within-subject variability; and increasing the number of replicates of observations per participants increases precision of only analytical variability. The sample sizes for biological variability studies can be planned better, with consideration of the primary estimate evaluated in a study and resource used to increase the precision of this estimate. Studies with sample sizes too small to achieve reasonable precision can be redeveloped or abandoned, minimizing research waste. If the desired precision of variability components is known, values for the sample size (number of participants, observations and replicates of observations) can be determined.

Interpretation of multi-analyte laboratory profiles, such as lupus anticoagulant (LAC) testing, requires integration of numerous test results and clinical information. However, this process is time-consuming, requires domain knowledge, and has limited scalability. Given that rule-based systems are rigid and difficult to maintain and large language models pose risks of "hallucinations," we developed and evaluated a novel retrieval-based approach to automate LAC profile interpretation to improve efficiency while maintaining fidelity and reliability for clinical use. Our method processes laboratory values through ensemble random forest (RF) models to retrieve the most similar expert interpretation from a curated database of 347 previously interpreted LAC profiles. The system's performance was assessed through technical accuracy metrics, a work flow efficiency timing study, and expert review of interpretation accuracy for held-out cases. Technical validation demonstrated high accuracy in retrieving correct interpretative elements (median: 93% accuracy, interquartile range [IQR]: 67% to 100%). Retrieval accuracy was highly correlated with the prevalence of a given element (&#x3c1; = 0.8307). In a timing study of 93 consecutive LAC profiles, ensemble-assisted interpretation achieved a median time of 22&#x2005;s per report (IQR: 16 s to 31&#x2005;s), representing a 78.6% total reduction compared to manual interpretation (median: 118&#x2005;s, IQR: 96 s to 172 s; t = 18.61, P < 0.001). Furthermore, 92.5% of AI-assisted interpretations were completed within 60&#x2005;s, exceeding the target of 85%. Expert review confirmed 100% concordance for LAC presence/absence. The proposed approach significantly improves efficiency of complex laboratory interpretation for LAC profiles, while maintaining consistency and high fidelity. Future studies will explore the method's inherent extensibility to the interpretation of other multi-analyte laboratory profiles.

The Africa Working Group, under the auspices of the Global Laboratory Quality Initiative of the Association for Diagnostics & Laboratory Medicine (ADLM), conducted several educational workshops in Africa to promote laboratory quality in the region. The challenges faced in maintaining quality in clinical laboratories in low-resource countries may differ from region to region and country to country. To better understand current gaps and inform our engagement with partners in Africa, a needs assessment survey was distributed to 1427 ADLM members in Africa and further disseminated through local partners, resulting in 121 responses from participants across 8 countries in the region. Additionally, interviews were conducted with laboratory professionals from the countries surveyed. The surveys primarily focused on quality management, continuing education, budgeting, infrastructure and funding, supplies, and staffing issues. The data collected from the surveys and interviews strongly indicate that respondents face major challenges in all areas assessed, including lack of funding, lack of equipment, lack of quality control (QC) and external quality assessment (EQA) materials, gaps in quality management, lack of appropriate budgeting, staffing issues, and lack of accessible training/continuing education. Consistent with the primary focus of ADLM;s Global Quality Initiative, improved access to continuing education was identified as the primary need by the participants (58%). The assessment highlights that while a strong role for educational workshops in mitigating some of the challenges continues to exist, there is also an opportunity to broaden the scope of the working groups beyond education.

This study assessed the ability of the complete blood count, differential, and reticulocyte (CDR) mode of the Mindray BC-6800 Plus analyzer to correct platelet aggregation by disaggregating platelets, evaluating how aggregation severity impacts accuracy to define an objective threshold for resampling. Agreement was assessed between CDR mode and the electrical impedance method for platelet counts in normal specimens using Pearson correlation, Passing-Bablok regression, and Bland-Altman analysis. The CDR mode's ability to correct false elevations caused by microcytes/fragments and false reductions caused by large platelets was evaluated by comparing CDR channel platelet count results (PLT-OF) with smear-estimated values (PLT-M) and fluorescence-method results (PLT-F). Disaggregation efficacy in aggregated samples was analyzed using the MC-80 digital morphology system. The optimal disaggregation threshold was determined by ROC analysis. In normal specimens, PLT-OF showed a significant correlation with the electrical impedance-method platelet count results (PLT-I) (r = 0.999, P < 0.001). Passing-Bablok regression (PLT-I = 1.00 &#xd7; PLT-OF) and Bland-Altman analysis demonstrated excellent agreement. For samples with PLT-I false abnormalities, the CDR mode effectively corrected both false elevations (due to microcytes/fragments) and false reductions (due to large platelets), showing no significant differences between PLT-OF and PLT-M or PLT-F (P > 0.05). In platelet-aggregated specimens, PLT-OF was significantly higher than PLT-I (P < 0.001). ROC analysis established 162 aggregates as the optimal threshold for effective disaggregation [area under the curve (AUC) = 0.776]. The dissociation rate was significantly greater in the mild vs severe-aggregation group (P = 0.004). Combining aggregate count with a PLT-OF to PLT-I difference (>115 &#xd7; 109/L) improved prediction accuracy (AUC = 0.841). The CDR channel of the Mindray BC-6800 Plus analyzer effectively corrects spurious platelet counts.

Our aim was to assess the knowledge, awareness, and confidence of US primary care providers (PCPs) in assessing multiple myeloma (MM) and related plasma cell disorders. A survey with quantitative and qualitative questions was conducted from October to December 2024. A total of 560 respondents met the criteria of a representative US PCP cohort, but only 300 respondents were invited to continue the rest of the survey. These respondents had ordered serum protein electrophoresis (SPEP) in the past 12 months and believed that SPEP should be ordered for plasma cell disorder diagnosis. Nearly half of the representative US PCP cohort (46%) do not order tests for plasma cell disorders. Among the 300 who do, there is generally good awareness of symptoms and high-risk characteristics, but only 28% follow guideline-recommended paired testing of SPEP and serum free light chains. The majority (80%) lack confidence in appropriate test selection and interpretation, often relying on peers and hematologists for guidance. Additionally, only 15% of the US PCPs have access to and use myeloma diagnostic ordering sets and panels, while 54% express interest in utilizing such tools. The PCPs identified barriers like insufficient education, collaboration, and specialist support to patient care and outcomes. The study reveals significant gaps in the understanding and practice of MM diagnostic testing in primary care. Results indicate insufficient knowledge and inadequate utilization of testing guidelines, including low confidence in test ordering and interpretation. The survey suggests a lack of comprehensive, guideline-adherent MM diagnostic panels offered by US laboratories.

The Duffy null variant (CC genotype) is associated with lower absolute neutrophil count (ANC) and increased risk of early discontinuation or ineligibility for certain treatments. Our primary objective was to evaluate the change in ANC over time among individuals on azathioprine or methotrexate. We also evaluated normality status (i.e., a designation of abnormal vs not) of ANC under genotype-adjusted and unadjusted thresholds. We used the UK Biobank to calculate genotype-adjusted and unadjusted thresholds. We included participants reporting azathioprine or methotrexate use in All of Us to examine the primary and secondary objectives. The primary outcome was the log-mean difference in ANC between 1 year before and 2 years post medication initiation. The secondary outcome was the odds of normality of ANC under genotype-adjusted vs unadjusted thresholds. We found that in azathioprine users, those with the CC genotype had nonstatistically significant lower ANC (log-mean difference, -0.017; 95% CI, -0.072 to 0.038). For methotrexate, those with the CC genotype had a statistically significant lower ANC (log-mean difference, -0.042; 95% CI, -0.074 to -0.009). Evaluations of unadjusted ANC thresholds in those on azathioprine revealed that CC individuals were more likely to be abnormal (odds ratio [OR], 7.51; 95% CI, 1.64 to 34.43). Findings were similar for methotrexate (OR, 4.35; 95% CI, 1.48 to 12.79). However, under Duffy-adjusted thresholds, there were no significant differences between both genotypes in either treatment group: azathioprine (P = .491) and methotrexate (P = .269). We conclude that Duffy-adjusted ANC reference ranges reduce the proportion of individuals classified as abnormal, with several implications for Duffy-null individuals.

High-sensitivity cardiac troponin assays are integral to diagnosing myocardial infarction. The current Roche Elecsys&#xae; Troponin T high-sensitivity Gen 5 assay (Gen 5) received FDA clearance in 2017. A sixth-generation assay was recently developed with improved analytical characteristics, including sensitivity and resistance to interferences. Reference ranges for this assay were reported, but analytical performance in the clinical laboratory has yet to be evaluated. The Elecsys&#xae; Troponin T high-sensitivity Gen 6 (Gen 6) assay was evaluated on the Roche cobas e 801 analyzers. Measurement limits of the assay were determined in lithium-heparin plasma specimens per CLSI EP17-A2. Repeatability and intermediate precision of QC materials and plasma specimens were determined according to CLSI EP05-A2 and EP15-A2, respectively. Serial dilutions of cardiac troponin T revealed that Gen 6 exhibits a positive bias relative to Gen 5 at concentrations >10&#x2005;ng/L. Below this point, the bias reverses, indicating concentration-dependent crossover. The limit of blank and limit of detection were 0.4&#x202f;ng/L and 1.2&#x202f;ng/L, respectively. The limit of quantitation was 2.2&#x2005;ng/L with a 10% CV. No cross-reactivity was observed on Gen 6 for skeletal muscle troponin T, cardiac troponin I, skeletal muscle troponin I, or troponin C at concentrations up to 10 000&#x202f;ng/L. Interference from hemolysis was not observed up to 5000&#x2005;mg/dL of hemoglobin. The Gen 6 assay demonstrates resistance to interference and greater analytical sensitivity, with a lower limit of detection and limit of quantitation relative to Gen 5, enabling broader coverage of healthy individuals.

Biochemical analysis of pericardial fluid (PF) can aid in the determination of the etiology of pericardial effusions. We validated 9 assays for albumin, amylase, blood urea nitrogen, cholesterol, direct and total bilirubin, lactate, lactate dehydrogenase, and total protein on the Roche Cobas c702 analyzer for PF. For each analyte, a recovery study was conducted to assess the effect of matrix; intra- and interassay precisions were determined at 3 levels; linearity was evaluated using 5 concentrations across the analytical measurement range; common interferences of hemoglobin, bilirubin, and triglycerides were investigated; and the stability at 22&#xb0;C up to 7 days, 2&#xb0;C to 8&#xb0;C up to 14 days, and -20&#xb0;C up to 4 weeks was studied. For all analytes, the recoveries were within 90% to 110%; the coefficient of variation ranged from 0% to 6% for intraassay and 1.1% to 6.9% for interassay precision. All assays exhibited linear response; the slopes ranged from 0.99 to 1.008, and R2 ranged from 0.999 to 1.000. No significant interferences of hemoglobin, bilirubin, or lipids were observed for all analytes except for direct bilirubin, which was negatively affected by hemoglobin and lipids. At -20&#xb0;C, all analytes were stable for 4 weeks. At 2&#xb0;C to 8&#xb0;C, all were stable for 14 days except lactate dehydrogenase (2 days) and direct bilirubin (2 days). At 22&#xb0;C, all were stable for 7 days except for direct bilirubin (8&#x2005;hours) and total bilirubin (3 days). No matrix effects were observed with the assays for these 9 analytes in PF on the Cobas c702 analyzer.

Mutations in MAPT, the tau gene, give rise to forms of frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17&#xa0;T), with abundant filamentous tau inclusions in brain cells. Some mutations that encode missense and deletion variants can give rise to a clinical picture of Pick's disease and filaments made of three-repeat tau in nerve cells. Here we report the electron cryo-microscopy (cryo-EM) structures of tau filaments from the brains of individuals with MAPT mutations D252V, G272V, S320F and &#x394;G389-I392. The two-layered Pick fold was present in the brains of individuals with mutations D252V and &#x394;G389-I392 who had also abundant tau inclusions in glial cells. By contrast, mutations G272V and S320F gave rise to a more open variant of the Pick fold, with residues 272-341 rotated by 20-25&#xb0; with respect to the rest of the structure. These findings show that missense mutations within the filament core can modify the Pick fold, generating closely related structural variants. In addition, we were able to reconstitute the Pick fold and some of its variants using seeded assembly with recombinant 0N3R tau carrying 12 serine or threonine to aspartate substitutions (PAD12) and missense mutations D252V, G272V and S320F. This work provides a foundation for the development of structure-based diagnostic and therapeutic approaches.

Hemolysis is a major cause of sample rejection in blood gas analysis. We aimed to develop a low-cost spectrometer for hemolysis detection in emergency laboratories. The spectrometer was developed using a complementary metal-oxide-semiconductor (CMOS) sensor and DVD-ROM grating. We evaluated the linearity, precision, and interference resistance of the instrument in accordance with CLSI and China National Accreditation Service for Conformity Assessment (CNAS) guidelines. In 51 clinical blood gas samples subjected to mechanically simulated hemolysis, the correlation between absorbance at 540&#x2005;nm (A540&#x2005;nm) and the hemolysis index (H index) determined by the Cobas c701 biochemical analyzer was evaluated. We utilized the hemolysis absorbance value that causes a maximum &#xb1;10% deviation in potassium (K+) and calcium ion (Ca2+) results as a reference to establish the rejection criteria for hemolysis in blood gas samples. The detection range of the spectrometer spanned 0.160< A540&#x2005;nm <2.67, exhibiting a highly significant correlation with the H index (P < 0.01). The between-run coefficient of variation (CV) for the device was below 5%, while the within-laboratory CV remained below 10%. At interfering substance levels of 8.0&#x2005;mg/dL (136.8&#x2005;&#x3bc;mol/L) for unconjugated bilirubin, 13.3&#x2005;mg/dL (226.7&#x2005;&#x3bc;mol/L) for conjugated bilirubin, and 431.4&#x2005;mg/dL (4.87&#x2005;mmol/L) for triglycerides, detection bias stayed within 10%. Hemolysis significantly altered pH, partial pressure of oxygen (pO2), K+, and Ca2+ (P < 0.05). By calculating the bias caused by hemolysis on the test results, rejection criteria were established: A540nm >0.671 hemolysis will affect K+ results and A540nm >0.960 hemolysis will affect Ca2+ results. This low-cost spectrometer enables reliable pre-analysis hemolysis detection, improving accuracy in blood gas testing.

Artificial intelligence (AI) is transforming the fields of genetics and genetic counseling, enhancing both clinical and laboratory practices. The rise of AI technologies has drawn attention to their potential impact on genetic counseling, particularly in patient diagnosis and the counseling processes. In the laboratory, AI plays a critical role in improving communication between laboratory genetic counselors and healthcare providers by automating routine tasks and optimizing workflows. These advancements allow genetic counselors to dedicate more time to addressing complex inquiries, improving genetic test selection, and helping providers interpret genetic test results. As AI continues to integrate into laboratory genetic counseling practice, it presents both opportunities and challenges. At the time of submission, there is a large knowledge gap regarding AI and its application to laboratory genetic counseling, given the lack of published information on this topic. This article summarizes existing literature, the history and current applications of AI in laboratory genetic counseling, examines its benefits and limitations, and explores future directions for its implementation in the field.

Carbamazepine is an antiepileptic and mood-stabilizing drug with complex pharmacokinetics and a narrow therapeutic index, making therapeutic drug monitoring (TDM) essential. The therapeutic reference range for carbamazepine at our laboratory is 20-40&#x2005;&#xb5;mol/L (4.7-9.4&#x2005;mg/L). Long-term laboratory data may help identify population trends, seasonal or demographic variation, and the suitability of patient-derived medians as a quality-assurance tool. This retrospective study analyzed 12 577 serum carbamazepine results obtained between May 2007 and December 2025 at Uppsala University Hospital. The total number of unique individuals providing the test results was 3147. Demographic variables (age, sex) and sampling dates were included. Yearly percentiles were used to evaluate long-term trends, and monthly variation was assessed for seasonal effects. Equivalence of analytical results was examined during the transition from the Architect ci8200 to the Cobas Pro c503 platform (February 2021). Annual test volume peaked in 2010 (n = 955) and declined to 160 by 2025. Despite reduced testing, carbamazepine concentrations increased steadily over time, with median values rising from 25.2&#x2005;&#xb5;mol/L (5.9&#x2005;mg/L) in 2007 to 31.2&#x2005;&#xb5;mol/L (7.4&#x2005;mg/L) in 2025. Seasonal analysis showed predictable drops in sampling during the Swedish summer vacation period but no meaningful variation in drug concentrations. The method transition in February 2021 showed strong agreement between platforms, with slightly higher values on the Cobas system (Cobas = 1.049 &#xd7; Architect - 0.164; R2 = 0.946). Carbamazepine concentrations in routine patient samples have gradually increased over 2 decades, independent of declining test frequency. Seasonal workload fluctuations did not affect observed levels. Patient-derived medians were stable and sensitive to method changes, supporting their role as complementary internal quality-control tools. Long-term TDM databases provide valuable insights for both clinical interpretation and laboratory quality assurance.

Early detection of monoclonal gammopathies is challenging, as they often remain undiagnosed until morbidity develops. Monoclonal proteins (M-proteins), particularly IgM, can interfere with the lipemia index (L-index) on chemistry analyzers, producing elevations in visually clear samples. The L-index may allow incidental detection of M-proteins, but its clinical value is uncertain. A retrospective chart review was conducted on 78 patients with visually clear serum samples with Siemens Atellica CH930 L-index &#x2265;2 to assess diagnoses and follow-up of previously known and newly identified M-proteins. L-index was measured on Atellica, Roche cobas Pro c503, and Ortho VITROS XT3400 in 40 additional samples from patients with an IgM M-protein to assess L-index sensitivity for M-proteins. A survey of laboratories examined current practices for managing such samples. Sixty-three of 78 patients with visually clear serum samples with a reproducible Atellica L-index &#x2265;2 had an M-protein(s) (34 previously known; 29 newly identified). Among these, IgM kappa predominated (47/63; 66%), and 5/29 newly identified cases were diagnosed with Waldenstr&#xf6;m macroglobulinemia. In 40 additional samples with known IgM M-proteins, only one had an elevated Atellica L-index, and none were elevated on cobas or Vitros. Survey results showed wide variation in managing these samples. An elevated L-index on the Atellica in visually clear serum samples, though insensitive, is predictive of the presence of an M-protein. These findings were inconsistent across platforms, highlighting the lack of L-index standardization. Protocol development of an elevated L-index in clear samples will help recognize and manage incidental M-proteins.

The soluble fms-like tyrosine kinase-1/ placental growth factor (sFlt-1/PlGF) ratio is widely used to aid in preeclampsia (PE) prognosis. Many laboratories apply cutoffs derived from the PRediction of short-term Outcome in preGNant wOmen with Suspected preeclampsIa Study (PROGNOSIS) using Elecsys results. However, differences between immunoassays and lack of international standardization introduce variability in sFlt-1/PlGF values and cutoffs, potentially affecting clinical interpretation. This study compared Elecsys and Kryptor immunoassays for sFlt-1 and PlGF and assessed their impact and the best cutoff for their ratio in PE risk classification. We compared Elecsys and Kryptor immunoassays in 160 samples from pregnant women with suspected PE. Correlation, receiver operating characteristic (ROC) performance (area under the curve [AUC]), and concordance across risk categories (low/intermediate/high) were assessed using Elecsys-based PROGNOSIS study cutoffs and a Kryptor-specific threshold. Kryptor yielded significantly lower PlGF and sFlt-1 concentrations than Elecsys (medians: 103.5&#x2005;ng/L vs 146.3&#x2005;ng/L and 3199&#x2005;ng/L vs 4034&#x2005;ng/L; both P < 0.001). Conversely, sFlt-1/PlGF ratios were higher with Kryptor (31 vs 25; P < 0.001). Diagnostic accuracy was comparable (AUC: Elecsys 0.809; Kryptor 0.810). Applying Elecsys cutoffs to Kryptor results shifted risk categorization, with Kryptor classifying fewer women as low risk (54.4%) compared with Elecsys (58.2%), and more women as high risk (24.3% vs 18.1%). Overall concordance was high (85%; &#x3ba; = 0.750), but among Elecsys-classified intermediate-risk samples, Kryptor agreed in only 63.2% and 31.6% were upgraded to high risk, reflecting Kryptor's tendency toward higher ratios. Elecsys and Kryptor show similar discriminative performance; however, immunoassay selection can alter PE risk stratification, particularly for intermediate risk cases. Method-specific thresholds are essential to avoid misclassification and optimize clinical decisions.

Excessive use of head computed tomography (CT) in mild traumatic brain injury (TBI) increases radiation exposure and contributes to emergency department crowding. Blood biomarkers glial fibrillary acid protein (GFAP) and ubiquitin carboxyl-terminal hydrolase L1 (UCH-L1) may improve risk stratification and guide clinical decision-making. We evaluated the diagnostic accuracy and prognostic value of a dual GFAP/UCH-L1 test, GFAP, and UCH-L1-individually and jointly-for detecting acute intracranial injury and predicting mortality. We conducted a retrospective observational study at a tertiary hospital from March 10, 2023, to March 31, 2025. Adult patients presenting within 12 h of mild TBI (Glasgow Coma Scale score 13 to 15) with available serum GFAP and UCH-L1 measurements were included. Biomarker concentrations were measured on the Alinity i. Dual GFAP/UCH-L1 test positivity was defined as GFAP &#x2265;35 pg/mL and/or UCH-L1 &#x2265;400 pg/mL. The primary outcome was acute intracranial injury on head CT. Diagnostic performance was assessed using CT as the reference standard. Secondary outcomes were 30- and 90-day mortality. A total of 2932 patients were included (median age 83 years; 57% women). Intracranial injury was identified in 230 patients (7.8%). The dual GFAP/UCH-L1 test showed high sensitivity (97.4%) and negative predictive value (98.6%). GFAP demonstrated better discrimination (area under the receiver operating characteristic [ROC] curve [AUC] 0.768) than UCH-L1 (AUC 0.642) and GFAP + UCH-L1 (0.742). In multivariable analysis, dual GFAP/UCH-L1 test positivity (odds ratio [OR] 7.28) independently predicted intracranial injury. UCH-L1 was associated with 30 day (hazard ratio [HR] 1.00028) and 90-day mortality (HR 1.00022). The dual GFAP/UCH-L1 test provides strong rule-out accuracy for acute intracranial injury, while UCH-L1 offers meaningful prognostic information. Biomarker guided assessment may reduce unnecessary CT use and support early risk stratification in emergency care.

Capillary self-collection (CSC) devices allow patients to collect blood samples at home, potentially reducing outpatient phlebotomy visits. This study aimed to (a) assess the patient experience with multiple commercially available CSC devices; (b) determine which laboratory tests are commonly ordered together, which could reduce the need for appointments; and (c) evaluate the analytical performance of these tests using CSC samples. User experience for 3 CSC devices was evaluated. Clinical feasibility was determined by comparing test results in paired sera collected by venipuncture (VP) and CSC devices. VP samples were centrifuged within 2&#x2005;h and tested immediately. CSC sera were centrifuged and tested both immediately and after delayed processing to simulate shipping temperature extremes (-20&#xb0;C or 40&#xb0;C). Basic metabolic panel, lipid panel, thyroid function cascade, and prostate-specific antigen were evaluated. Differences between VP and CSC collections were characterized according to their statistical and clinical differences. Patients reported CSC devices were easy to use and painless. Clinically significant differences between VP and CSC sera processed immediately were limited to potassium and bicarbonate. Following delayed processing of CSC serum samples (48&#x2005;h at room temperature), clinically significant differences in potassium, bicarbonate, and glucose were observed. Frozen samples could not be analyzed. A 48&#x2005;h delay at 40&#xb0;C caused clinically significant differences in all analytes except creatinine, lipid panel, prostate-specific antigen, and the thyroid cascade. Potassium, bicarbonate, and glucose were not stable at room temperature in CSC sera. CSC serum specimens may be a viable option but require analyte-specific evaluation and consideration of transportation conditions.

Intrahepatic cholestasis of pregnancy (ICP) requires timely diagnosis to guide management, including initiation of ursodeoxycholic acid (UDCA) and delivery planning. LC-MS/MS provides fractionated bile acid profiles but has a turnaround time (TAT) of approximately 1 week, delaying treatment. Enzymatic total bile acid (TBA) assays offer faster results but measure all bile acids collectively, which some have reported can be impacted by UDCA treatment. We validated the Diazyme enzymatic TBA assay on a Roche Cobas analyzer, assessing analytical measurement range, reference interval, precision, and accuracy vs LC-MS/MS. The comparison cohort included samples from 100 pregnant patients (median age 31 years; gestational age 34 weeks) presenting with pruritus and suspected of ICP. A secondary analysis examined UDCA and tauroursodeoxycholic acid in a UDCA-detectable subset. TAT was evaluated retrospectively in 971 specimens. We verified the assay's analytical measurement range of 1 to 180&#x2005;&#xb5;mol/L and reference interval of <10&#x2005;&#xb5;mol/L. Precision was <10%, and method comparison revealed minimal bias, with 98% clinical concordance. In the UDCA subset (n = 19), UDCA and tauroursodeoxycholic acid represented minor fractions of total TBA. Operationally, the enzymatic assay provided rapid results, with a median TAT of 5.6&#x2005;h from collection (range 0.6-36.8) and 0.5&#x2005;h from receipt (range 0.2-22.3); delays were mainly due to transport. The Diazyme enzymatic TBA assay on Roche Cobas demonstrates robust performance and rapid turnaround, supporting timely ICP diagnosis and management. LC-MS/MS remains useful for fractionated profiling but is limited by longer TAT.

Glial fibrillary acidic protein (GFAP) and neurofilament light chain (NfL) are critical blood-based biomarkers for the diagnosis and monitoring of a wide range of central nervous system (CNS) injuries. Current plasma-based assays require centrifugation, which prolongs turnaround time (TAT) and limits utility in emergency settings. This study aimed to develop and validate a rapid, automated chemiluminescence immunoassay for the direct quantification of GFAP and NfL in whole blood. We used a dual-antibody sandwich chemiluminescence immunoassay to measure GFAP and NfL. The assay was optimized for whole blood through adjustments of reagent formulation and implementation of a hematocrit (HCT) correction algorithm. Performance was evaluated in terms of precision, linearity, limit of quantification, and interference. In total, 115 paired whole blood and plasma samples from 78 acute stroke patients were analyzed to assess diagnostic accuracy. The HCT correction algorithm effectively minimized measurement bias across a wide HCT range (35%-55%). Precision of GFAP and NfL in whole blood and plasma showed coefficients of variation below 5% across a broad concentration range. Excellent correlations were observed between whole blood (HCT-corrected) and plasma measurements for both GFAP (R2 = 0.990) and NfL (R2 = 0.955). Diagnostic accuracy for distinguishing hemorrhagic from ischemic stroke was comparable between whole blood and plasma (AUC 0.86 vs 0.84, P = 0.58) in this population. The novel GFAP and NfL whole-blood assay, due to its shorter TAT, high consistency with plasma, and broad analytical range, demonstrated the potential to accelerate stroke triage in acute care settings.

Per CLSI EP46 (2025), the validation of a test device, when compared to a reference device, should focus on evaluating the total analytical error (TAE) and show that it is within an allowable total error (ATE). Specifically, FDA Assay criteria (2013) suggest that 95% of test device data should be within ATE from those of a reference device. This article presents statistical tools for assessing agreement in method comparisons and assay validations that satisfy the above guidance. Here, we emphasize the concept and provide examples using a web tool. The validated web tool, free for public use, is available to satisfy the above hard-defined criteria for device acceptance and provides information regarding accuracy and precision. We use test and reference measurements of selected clinical chemistry and hematology analytes as examples. We selected 4 analyte cases to evaluate: precise and accurate, precise and inaccurate, imprecise and accurate, and imprecise and inaccurate. Among the 4 analytes, the precise and accurate case is the only one that passed the FDA Assay criteria. We also present the misleading results indicated by traditional paired t-test and regression analyses, which rejected the case that passed the FDA Assay criteria. The freely accessible and validated web tool successfully identifies analytes that meet the FDA Assay criteria and provides an indication of when an investigation of a potential precision and/or accuracy problem is required.