Impact of Academia-Government Collaboration on Laboratory Medicine Standardization in South Korea: analysis of eight years creatinine proficiency testing experience.

OBJECTIVES: To evaluate the performance of the Academia-Government Collaboration for Laboratory Medicine Standardization in Korea (KR-STDZN) based on data from KR-STDZN proficiency testing (KR-STDZN-PT) for creatinine over eight years (2015-2022). METHODS: We used KR-STDZN-PT data of creatinine tests from 2015 to 2022. Acceptance of the participating institutions' test results was assessed by calculating the acceptance performance as absolute bias (absBias%), total coefficient of variance (tCV%), and total error (TE%) for each sample using six measurements from each institution and true values of each reference material. The test result was considered acceptable when absBias%, tCV%, and TE% were <5.10, <3.20, and <11.40 %, respectively. The proportion of acceptable institutions among all participating institutions in each round was defined as the acceptance rate. Improvements in absBias%, tCV%, and TE% were analyzed using creatinine concentration ranges in samples. RESULTS: The number of participating institutions increased from 2015 to 2017 but remained consistent since 2018. The acceptance rates for absBias% and TE% increased from 52.2 and 77.6 %, in 2015 and to 90.7 and 96.3 %, in 2022, respectively. The acceptance rate for tCV% remained in the 90 % range for eight years. When creatinine <3 mg/dL, mean absBias%, and mean TE% improved significantly in 2021-2022 compared to 2015-2016 (p<0.05). When creatinine >3 mg/dL, acceptance performance did not improve. Mean tCV% remained consistent annually regardless of creatinine concentration. No significant variations in test methods were observed. CONCLUSIONS: The collaboration between academia and the government improved creatinine testing quality. Nevertheless, KR-STDZN must be expanded and refined.

Usefulness and performance evaluation of serum KL-6 and SP-A assays in healthy individuals and patients with interstitial lung disease.

BACKGROUND: Interstitial lung abnormalities (ILAs) are associated with the risk of progression to interstitial lung diseases (ILDs). Krebs von den Lungen 6 (KL-6) and surfactant protein (SP)-A have been used as biomarkers of ILDs. In this study, we evaluated the levels of these biomarkers and identified their clinical correlations in healthy individuals to assess their usefulness in the diagnosis of ILAs. METHODS: The patient samples were categorized into three groups: healthy, disease, and ILD groups. We used the automated immunoassay HISCL KL-6 and SP-A assay kits. The analytical performance evaluation involved precision, linearity, comparison, establishment of reference intervals, and determination of the cutoff points. We also analyzed the correlations between presence of abnormalities on chest radiography and computed tomography (CT) or pulmonary function test (PFT) and serum levels in the healthy group. RESULTS: KL-6 and SP-A assays showed good analytical performance. The KL-6 and SP-A cutoff values were 304 U/mL and 43.5 ng/mL between the ILD and healthy groups, respectively, which were lower than the values recommended by the manufacturer. In the clinical correlations with radiological findings, SP-A values in subjects with lung abnormalities on CT scans were significantly higher than those in normal scans. There was no significant difference in KL-6 and SP-A levels among PFT patterns; however, both serum levels in the mixed pattern showed higher values than those in the other patterns. CONCLUSIONS: The results revealed a positive association between increased serum levels of SP-A and KL-6 and clinical characteristics as incidental findings on chest imaging and reduced lung function.

A New Strategy for Evaluating the Quality of Laboratory Results for Big Data Research: Using External Quality Assessment Survey Data (2010-2020).

Background: To ensure valid results of big data research in the medical field, the input laboratory results need to be of high quality. We aimed to establish a strategy for evaluating the quality of laboratory results suitable for big data research. Methods: We used Korean Association of External Quality Assessment Service (KEQAS) data to retrospectively review multicenter data. Seven measurands were analyzed using commutable materials: HbA1c, creatinine (Cr), total cholesterol (TC), triglyceride (TG), alpha-fetoprotein (AFP), prostate-specific antigen (PSA), and cardiac troponin I (cTnI). These were classified into three groups based on their standardization or harmonization status. HbA1c, Cr, TC, TG, and AFP were analyzed with respect to peer group values. PSA and cTnI were analyzed in separate peer groups according to the calibrator type and manufacturer, respectively. The acceptance rate and absolute percentage bias at the medical decision level were calculated based on biological variation criteria. Results: The acceptance rate (22.5%-100%) varied greatly among the test items, and the mean percentage biases were 0.6%-5.6%, 1.0%-9.6%, and 1.6%-11.3% for all items that satisfied optimum, desirable, and minimum criteria, respectively. Conclusions: The acceptance rate of participants and their external quality assessment (EQA) results exhibited statistically significant differences according to the quality grade for each criterion. Even when they passed the EQA standards, the test results did not guarantee the quality requirements for big data. We suggest that the KEQAS classification can serve as a guide for building big data.

Total PSA Test Performance Evaluation on Cobas Pure and Measurement System Comparison with Different Calibrators.

BACKGROUND: This study investigated the performance evaluation of total prostate specific antigen (tPSA) testing using Cobas Pure integrated solutions system (calibrated against WHO IS 96/670) and the comparison with established measurement systems with different traceability. METHODS: The evaluation was performed in terms of imprecision, linearity, detection limit, and correlation with Alinity i (calibrated against WHO IS 96/670) and Unicel DxI 800 (calibrated against the manufacturer's working calibrators). RESULTS: Within-laboratory reproducibility and repeatability were observed less than 1.2%. Linearity was achieved within the claimed analytical measurement range. The claimed LoB and LoD were experimentally verified. All the correlation coefficients among the assays indicated good correlation, but the significant mean bias with Unicel DxI 800 using a different calibrator were observed. CONCLUSIONS: Since the tPSA calibrators against different traceability is still commercially available, our research could convey the impact of calibration on tPSA results as well as the performance information of a new assay for tPSA.

Accuracy of the New Creatinine-based Equations for Estimating Glomerular Filtration Rate in Koreans.

Background: New creatinine-based estimated glomerular filtration rate (eGFR) equations, including the 2021 Chronic Kidney Disease Epidemiology Collaboration (2021 CKD-EPI) and European Kidney Function Consortium (EKFC) equations, have been introduced recently. We assessed the performance of the 2021 CKD-EPI and EKFC equations in the Korean population. Methods: We analyzed 1,654 Korean patients aged ≥18 years who underwent chromium-51-ethylenediamine tetraacetic acid GFR measurements (mGFR). Bias (eGFR-mGFR), root mean square error (RMSE), and proportion of eGFR within 30% of mGFR (P30) of the 2009 CKD-EPI, 2021 CKD-EPI, and EFKC equations were compared. The concordance rate between eGFR and mGFR categories was evaluated. Both eGFR and mGFR categories were classified into six groups: ≥90, 89-60, 59-45, 44-30, 29-15, and <15 mL/min/1.73 m2. Results: The median bias (mL/min/1.73 m2) was 1.8 for the 2009 CKD-EPI equation, 4.8 for the 2021 CKD-EPI equation, and -0.3 for the EKFC equation. The P30 and RMSE were 78.2% and 17.0 for the 2009 CKD-EPI equation, 75.6% and 17.4 for the 2021 CKD-EPI equation, and 80.0% and 16.7 for the EKFC equation, respectively. The overall GFR category concordance rate between eGFR and mGFR was 63.4% for the 2009 CKD-EPI equation, 60.5% for the 2021 CKD-EPI equation, and 61.0% for the EKFC equation. Conclusions: Among the three eGFR equations, the EKFC equation had the smallest bias and highest P30 in Koreans. The 2009 CKD-EPI equation had a lower bias than the 2021 CKD-EPI equation.

Association between measured or calculated small dense low-density lipoprotein cholesterol and oxidized low-density lipoprotein in subjects with or without type 2 diabetes mellitus.

BACKGROUND: Small dense low-density lipoprotein (sdLDL) possesses atherogenic potential and is predicted to be susceptible to atherogenic modifications, which further increases its atherogenicity. However, studies on the association between measured or estimated sdLDL cholesterol (sdLDL-C) levels and atherogenic modification in diverse population groups are lacking. METHODS: Surplus serum samples were collected from male subjects with type 2 diabetes mellitus (DM) under treatment (n = 300) and without DM (non-DM; n = 150). sdLDL and oxidized LDL (oxLDL) levels were measured using the Lipoprint LDL subfractions kit (Quantimetrix Corporation) and the Mercodia oxidized LDL competitive enzyme-linked immunosorbent assay kit (Mercodia), respectively. The estimated sdLDL-Cs were calculated from two relevant equations. The effects of sdLDL-C on oxLDL were assessed using multiple linear regression (MLR) models. RESULTS: The mean (±SD) of measured sdLDL-C and oxLDL concentrations were 11.8 ± 10.0 mg/dl and 53.4 ± 14.2 U/L in the non-DM group and 0.20 ± 0.81 mg/dl and 46.0 ± 15.3 U/L in the DM group, respectively. The effects of measured sdLDL-Cs were significant (p = 0.031), whereas those of estimated sdLDL-Cs were not (p = 0.060, p = 0.116) in the non-DM group in the MLR models. The effects of sdLDL-Cs in the DM group were not significant. CONCLUSION: In the general population, high level of sdLDL-C appeared to be associated with high level of oxLDL. The equation for estimating sdLDL-C developed from a general population should be applied with caution to a special population, such as patients with DM on treatment.

Intuitive Modification of the Friedewald Formula for Calculation of LDL-Cholesterol.

Background: High LDL-cholesterol (LDL-C) is an established risk factor for cardiovascular disease and is considered an important therapeutic target. It can be measured directly or calculated from the results of other lipid tests. The Friedewald formula is the most widely used formula for calculating LDL-C. We modified the Friedewald formula for a more accurate and practical estimation of LDL-C. Methods: Datasets, including measured triglyceride, total cholesterol, HDL-cholesterol, and LDL-C concentrations were collected and assigned to derivation and validation sets. The datasets were further divided into five groups based on triglyceride concentrations. In the modified formula, LDL-C was defined as total cholesterol - HDL-cholesterol - (triglyceride/adjustment factor). For each group, the adjustment factor that minimized the difference between measured LDL-C and calculated LDL-C using modified formula was obtained. For validation, measured LDL-C and LDL-C calculated using the modified formula (LDL-CM), Friedewald formula (LDL-CF), Martin-Hopkins formula (LDL-CMa), and Sampson formula (LDL-CS) were compared. Results: In the derivation set, the adjustment factors were 4.7, 5.9, 6.3, and 6.4 for the groups with triglyceride concentrations <100, 101-200, 201-300, and >300 mg/dL, respectively. In the validation set, the coefficient of determination (R2) between measured and calculated LDL-C was higher for LDL-CM than for LDL-CF (R2=0.9330 vs. 0.9206). The agreement according to the National Cholesterol Education Program Adult Treatment Panel III classification of LDL-C was 86.36%, 86.08%, 86.82%, and 86.15% for LDL-CM, LDL-CF, LDL-CMa, and LDL-CS, respectively. Conclusions: We proposed a practical, improved LDL-C calculation formula by applying different factors depending on the triglyceride concentration.

Strategies to shorten turnaround time in outpatient laboratory.

BACKGROUND: Turnaround time (TAT) is one of the most important indicators of laboratory quality. For the outpatient routine chemistry tests whose results are checked by clinicians on the same day, we set a quality goal that >90% of these samples should be reported within 60 min. As more than 20% of the samples failed to achieve this goal in 2020, we introduced an additional autoanalyzer and a real-time monitoring system to improve this rate. METHODS: As the TAT of the pre-analytical phase is the greatest contributor to TAT, we divided it into sampling, sample transport, and sample preparation times. An additional autoanalyzer was introduced, and its effect on TAT improvement was evaluated with the TAT data of June and July 2020. A real-time monitoring system was introduced to sort delayed samples, and its effect was assessed with the TAT data of June and July 2021. TAT data from December 2019 to January 2020 were set as baseline controls. RESULTS: The preparation time comprised the largest proportion of TAT. Although there was a slight decrease in overall TAT after the introduction of the above two strategies, the target TAT achievement rate increased significantly from 78.5% to 88.7% (p < 0.001). CONCLUSIONS: We checked the cause of TAT prolongation and introduced new strategies to improve it. The addition of an autoanalyzer per se was not so effective but was better when combined with the real-time monitoring system. Such strategies would increase the quality of the laboratory services.

Practical Considerations for Clinical Laboratories in Top-down Approach for Assessing the Measurement Uncertainty of Clinical Chemistry Analytes.

Background: The top-down (TD) approach using internal quality control (IQC) data is regarded a practical method for estimating measurement uncertainty (MU) in clinical laboratories. We estimated the MU of 14 clinical chemistry analytes using the TD approach and evaluated the effect of lot changes on the MU. Methods: MU values were estimated using subgrouping by reagent lot changes or using the data as a whole, and both methods were compared. Reagent lot change was simulated using randomly generated data, and the mean values and MU for two IQC datasets (different QC material lots) were compared using statistical methods. Results: All MU values calculated using subgrouping were lower than the total values; however, the average differences were minimal. The simulation showed that the greater the increase in the extent of the average shift, the larger the difference in MU. In IQC data comparison, the mean values and MU exhibited statistically significant differences for most analytes. The MU calculation methods gave rise to minimal differences, suggesting that IQC data in clinical laboratories show no significant shift. However, the simulation results demonstrated that notable differences in the MU can arise from significant variations in IQC results before and after a reagent lot change. Additionally, IQC material lots should be treated separately when IQC data are collected for MU estimation. Conclusions: Lot changes in IQC data are a key factor affecting MU estimation and should not be overlooked during MU estimation.

Age Group-specific Reference Intervals for the Elecsys Anti-Müllerian Hormone Assay in Healthy Korean Women: a Nationwide Population-based Study.

Background: Anti-Müllerian hormone (AMH) is one of the most reliable markers of ovarian reserve. Automated AMH assays are widely used in clinical laboratories, but reference intervals for the Elecsys AMH assay for Asian populations have not yet been determined. We aimed to determine reference intervals in healthy Korean women. Methods: The study included 1,450 women aged 19 to 54 years who participated in the Korea National Health and Nutrition Examination Survey between 2013 and 2016. The study participants were divided into seven 5-year age groups. AMH and progesterone concentrations were measured using Roche Elecsys assays, and bone morphogenetic protein-15 (BMP15) was genotyped for the detection of major variants. Age group-specific reference intervals for AMH were established as recommended by the CLSI EP28-A3c guidelines. Results: The mean age was 37.4 years. AMH concentrations decreased with increasing age, especially after 40 years, with the median AMH decreasing from 30.9 pmol/L in participants of 19-24 years to 0.071 pmol/L in participants of 50-54 years. The mid-95 percentile AMH reference intervals decreased from 7.93-81.21 pmol/L in participants of 19-24 years to 0.07-3.86 pmol/L in participants of 50-54 years. Disease-associated BMP15 variants were not detected. Conclusions: We determined Elecsys AMH assay reference intervals in healthy Korean women. The results may provide basic information for the interpretation of AMH concentrations and assessment of ovarian reserve in Korean women.

Evaluation of Intra- and Interlaboratory Variations in SARS-CoV-2 Real-Time RT-PCR Through Nationwide Proficiency Testing.

OBJECTIVE: This study aimed to examine the intra- and interlaboratory variations of cycle threshold (Ct) values using the nationwide proficiency testing for SARS-CoV-2. METHODS: Triplicated strong-positive contrived samples duplicated weak-positive contrived samples, and 2 negative samples were transported to participating laboratories in October 2021. RESULTS: A total of 232 laboratories responded. All except 4 laboratories correctly answered. Six false-negative results, including 2 false-negatives with Ct values beyond the threshold and 1 clerical error, were noted from weak-positive samples. Intralaboratory variations of Ct values of weak-positive and strong-positive samples were not acceptable (Ct > 1.66) in 17 and 7 laboratories, respectively. High interlaboratory variations of Ct values (up to 7 cycles) for the 2 commonly used polymerase chain reaction (PCR) reagents were observed. CONCLUSION: The overall qualitative performance was acceptable; intralaboratory variation was acceptable. However, interlaboratory variations of Ct values were remarkable even when the same PCR reagents were used.

Effect of Outlier Elimination on the 99th Percentile Upper Reference Limits of High-Sensitivity Cardiac Troponin I Assays Based on a Strictly Selected Healthy Reference Population.

BACKGROUND: We established high-sensitivity cardiac troponin I (hsTnI) 99th percentile upper reference limits (URLs) for the Centaur XPT High-Sensitivity Troponin I assay (Centaur hsTnI; Siemens, Erlangen, Germany) and Atellica IM High-Sensitivity Troponin I assay (Atellica hsTnI; Siemens) and assessed the effect of outlier elimination. METHODS: The reference population comprised 380 men and 387 women, satisfying the strict systematic reference population criteria. After reference population verification by the N-terminal pro-B-type natriuretic peptide (NT-proBNP) assay, 99th percentile URLs for Centaur hsTnI and Atellica hsTnI were calculated before and after outlier elimination. RESULTS: The 99th percentile URL for Centaur hsTnI was 60.4 (men, 74.7; women, 57.5) ng/L and that for Atellica hsTnI was 59.6 (men, 75.2; women, 55.1) ng/L. After the elimination of 61 (8.0%) outlier samples in Centaur hsTnI and 58 (7.6%) in Atellica hsTnI, the 99th percentile URLs were 13.5 ng/L (men, 15.3 ng/L; women, 11.9 ng/L) and 13.4 ng/L (men, 15.5 ng/L; women, 12.9 ng/L), respectively, significantly lower than those before outlier elimination. The CVs at the 99th percentile URLs were 5.2% and 3.5%, respectively. The measurable fractions among the reference population were 91.5% and 93.4%, respectively. Performance evaluation of Atellica B-type natriuretic peptide (BNP), Atellica NT-proBNP, Centaur hsTnI, and Atellica hsTnI showed outstanding results. CONCLUSIONS: The Korean hsTnI 99th percentile URLs calculated in this study were significantly lower after outlier elimination than before. Centaur hsTnI and Atellica hsTnI meet the "Guideline acceptable" and "Level 3 (second generation, high sensitivity)" requirements, satisfying international standards.

Semiquantitative Analysis of the Results of an Image-Based Urine Sediment Analyzer Considering the Preanalytical Factors.

BACKGROUND: Automated microscopic platforms are increasingly used in clinical laboratories for rapid analysis of samples. However, it is important to present the results quantitatively or semiquantitatively because automated platforms use various technologies for analysis as well as different sediment preparation methods. The results of cell counting using an on screen image review program for the cobas u 701 analyzer (Roche Diagnostics Interna-tional, Rotkreuz, Switzerland) differed from those obtained by manual microscopic examination (MME). This study was performed to investigate the difference of results among analyzer, on-screen image review and MME. METHODS: Freshly collected urine specimens from outpatients were used. We calculated the mean, standard deviation, and 95% confidence interval for red and white blood cell (RBC/WBC) quantitative results obtained using the cobas u 701 analyzer. These results were compared to those obtained by manual counting. RBC and WBC counts determined with the cobas u 701 analyzer were compared to those obtained by MME per unit field. RESULTS: The semiquantitative results of MME were graded as 0 - 2, 3 - 5, 6 - 10, 11 - 20, 21 - 30, and many or numerous cells/high power field (HPF). The RBC and WBC counts determined by image analyses showed the tendency to be one grade higher than those from MME in the range of 3 to 5/HPF to many/HPF. The results of nearly all samples with 0 - 2/HPF and numerous/HPF for RBC and WBC counts were consistent with the grade found by MME. CONCLUSIONS: The one-grade difference may have been caused by the differences of preanalytical factors in the sample volume, centrifugal force, urine concentration ratio, or sediment volume/area of the slide. When reporting the results of image analyses, RBC and WBC counts should be raised by one grade to compensate for MME. Each laboratory needs to verify the on-screen review of images corresponding to the microscopic field of view according to the clinical laboratory's specific preanalytical practices.

Status of Four Trace Elements in Elderly Patients with Online Hemodiafiltration.

BACKGROUND: Essential trace elements play key roles in multiple biological systems, and hemodialysis patients are at risk for deficiency of essential trace elements. The aim of the study was to assess the essential element status in end stage renal disease patients undergoing online hemodiafiltration (online HDF) in outpatient dialysis clinic. METHODS: A total of 28 Korean patients with regular online HDF were included. Blood samples were collected before and after one HDF session, and serum concentrations of zinc, copper, selenium, and manganese were simulta-neously measured by inductively coupled plasma mass spectrometry. RESULTS: Selenium, zinc, copper deficiencies were observed in 71.4%, 35.8%, and 21.4%, compared with the reference range. No patients revealed manganese deficiency. After the HDF, the post-HDF level significantly increased in all trace elements, compared with the pre-HDF (11.2% for selenium, 10.7% for copper, and 6.6% for zinc). However, 50% patients were still deficient for selenium at the post-HDF. CONCLUSIONS: Our data suggest that the patients undergoing online HDF are at an increased risk of trace element deficiency, especially for selenium.

Identification of a Splice Variant (c.5074+3A>C) of BRCA1 by RNA Sequencing and TOPO Cloning.

Grading the pathogenicity of BRCA1/2 variants has great clinical importance in patient treatment as well as in the prevention and screening of hereditary breast and ovarian cancer (HBOC). For accurate evaluation, confirming the splicing effect of a possible splice site variant is crucial. We report a significant splicing variant (c.5074+3A>C) in BRCA1 in a patient with recurrent ovarian cancer. Next-generation sequencing (NGS) of BRCA1/2 from patient's peripheral blood identified the variant, which was strongly suspected of being a splicing mutation based on in silico predictions. Direct RNA analysis yielded multiple transcripts, and TOPO cloning of the complementary DNA (cDNA) and Sanger sequencing revealed an aberrant transcript with an insertion of the first 153 bp of intron 17, and another transcript with the 153 bp insertion along with an exon 18 deletion. A premature termination codon was presumed to be formed by the 153 bp partial intron retention common to the two transcripts. Therefore, BRCA1 c.5074+3A>C was classified as a likely pathogenic variant. Our findings show that active use of functional studies of variants suspected of altered splicing are of great help in classifying them.

Development and validation of a novel sepsis biomarker based on amino acid profiling.

BACKGROUND & AIMS: Sepsis is a potentially fatal condition influenced by pathogens and host factors. Current sepsis biomarkers such as white blood cell count and C-reactive protein and procalcitonin levels show unsatisfactory performance in terms of diagnostic sensitivity and specificity in clinical practice. Thus, we developed and validated a new sepsis biomarker based on amino acid profiling. METHODS: We used two independent groups. The training and validation groups included 161 and 22 healthy controls, 123 and 50 patients with systemic inflammatory response syndrome, and 115 and 45 patients with sepsis, respectively. Using mass spectrometry, we measured and analyzed serum amino acid levels to select candidate amino acids that could differentiate sepsis from other conditions. Then, several possible multivariate indexes were developed by generating formulae with different combinations of candidate amino acids. The formula showing the best performance was selected and validated further. RESULTS: Kynurenine, tryptophan, phenylalanine, arginine, aspartic acid, glutamic acid, and glutamine were selected as candidate amino acids. Ten possible formulae were generated, and the formula with the highest diagnostic performance, which included kynurenine, tryptophan, phenylalanine, and arginine, was selected. In the validation group, the area under the receiving operating characteristic curve of the selected multivariate index (0.931) was similar to that of procalcitonin (0.945). Moreover, the generated multivariate index showed potential as a prognostic marker. CONCLUSIONS: Serum amino acid composition in patients with sepsis differs significantly from that in healthy individuals and patients with inflammation only. The newly developed multivariate index is expected to be implementable as a sepsis biomarker in clinical practice in the near future.