bluuebunny/crossref_metadata_2025 huggingface dataset:
A-144 Liquid chromatography multiple reaction monitoring-based assay for quantitative assessment of fibrinogen levels in clinical samples
Authors: Chavan S , Bokka R , Venkataraman S , Peterson J , Tange J , Leger R , Seheult J , Pruthi R , Chen D , Pandey A
Published: October 2024

Background

Fibrinogen plays a critical role in hemostasis and is essential to clot formation. Changes in the quantity or activity of fibrinogen can lead to disorders affecting coagulation. Thus, it is essential to precisely quantify fibrinogen levels to differentiate between hyperfibrinogenemia in hypercoagulable disorders and dysfibrinogenemia, hypo/afibrinogenemia, and irregular bleeding. Current diagnostic tests that use antibodies to detect fibrinogen or activity-based measures are susceptible to interference from heparin contamination, hemolysis, and hyperbilirubinemia. Here, we evaluated the use of targeted proteomics for developing a quantitative assay for fibrinogen in plasma samples.

Methods

Four peptides each for alpha and gamma chains of fibrinogen were synthesized using FMOC chemistry on a Liberty Blue (CEM Corp) peptide synthesizer. A targeted multiple reaction monitoring (MRM) assay was developed and optimized using Dionex Ultimate 3000 HPLC connected to a TSQ Altis triple quadrupole mass spectrometer (Thermo Scientific). Evaluations were conducted using a range of standards, such as pure protein, WHO plasma and commercially available plasma with known fibrinogen amounts for the accurate quantitation of fibrinogen. Calibration curves were generated using different dilutions in various matrices. The repeatability and precision of the assay were evaluated across five different days. The plasma samples were processed and digested in a 96-well plate and subjected to C18 clean-up using a Bravo automated liquid handler (Agilent). All samples were spiked-in with corresponding heavy-labeled peptides prior to digestion with trypsin. Multiple reaction monitoring assays were performed on tryptic digests of plasma samples.

Results

Calibration curves using commercial plasma for all peptides showed linearity from 20 mg/dl to 1,308 mg/dl. This covers the entire clinical range including normal (200-350 mg/dl), deficient states (<100 mg/dl) and excess production (>350 mg/dl) that can be used for evaluation of bleeding disorders including consumption coagulopathy and hyperfibrinogenemia. Based on the overall performance in terms of linearity, limit of detection/quantitation, carryover and coefficient of variation (CV) across five different days, two peptides per chain were finally selected for developing the quantitative assay. The intra-assay variability was <5% and intra-day CV was ∼20%. This assay is currently being tested on a variety of clinical samples representing a range of fibrinogen levels.

Conclusions

Our study focused on assessing whether multiplexed MRM assay could be implemented to measure fibrinogen levels in plasma. Our data indicate the feasibility of MRM-based assays for determining fibrinogen levels as an alternative to existing non-mass spectrometry-based activity or antigen assays.
A-149 Comparison of Mass Spectrometry to Sickle Solubility for Confirmation of Low Percentages of Hemoglobin S
Authors: Olson T , Rangan A , Kandler R , Pappas A , Yang B , Zhang X , Herrick J L
Published: October 2024

Background

Hemoglobin S (HbS) causes sickling disorders. In the laboratory, confirmation of HbS in patient samples with varying percentages is essential. Assessment is routinely performed using capillary electrophoresis (CE) and/or high-performance liquid chromatography. Sickle solubility (SS) exploits decreased solubility of HbS under reducing conditions and is a confirmatory test. SS is unreliable with low HbS, elevated HbF, severe anemia, erythrocytosis, hyperglobulinemia, extreme leukocytosis, hyperlipidemia, or increased serum proteins. Infant samples commonly have low percentages of HbS, high HbF, and limited sample volume. Intact globin chain Mass Spectrometry (MS) is used to identify abnormal hemoglobin variants by detecting the mass change (normal α/β/γ mass =15127/15867/15996 amu), respectively). MS has potential advantages in sensitivity and efficiency over SS and requires lower input sample volume. Herein we compare the utility of MS versus SS for confirmation of HbS.

Methods

Twenty-three EDTA-anticoagulated blood samples containing low percentages of HbS were identified. SS was performed on 30μL washed packed cells (requires 200μL whole blood) per standard methods. Intact Quadrupole Time-of-Flight MS (Accurate-Mass Q-TOF LC/MS 6520, Agilent Technologies, California) was performed using 10μL whole blood sample in acetonitrile solution reagent and results were compared. HbS and HbF levels were assayed by capillary electrophoresis (Sebia, France) per standard methods.

Results

Select results are summarized in Table 1. SS detected HbS percentages to 4.8% when HbF was low but not 6% HbS when Hb F is high. MS showed no HbF interference and identified HbS peaks (at 15837 amu) down to 3.1%. HbS percentages <3.1 were undetected by both methods.

Conclusions

Overall, MS was superior to SS for confirming HbS, particularly in samples with low variant percentage and/or elevated HbF. MS is also optimal when sample volume is minimal, such as for infants. Percentages of HbS <3% require methods such as IEF or DNA sequencing for confirmation.
A-154 Non-standard body fluid validation of d-dimer analysis of organox liver perfusate to support clinical evaluations of donor livers prior to transplantation
Authors: Koster H , Majkut N , Krum K , Maggiore J , Gant Kanegusuku A
Published: October 2024

Background

Non-standard body fluid analysis is considered a challenge to laboratories since very few have been approved by the FDA for routine testing. As medical procedures evolve, the laboratory must balance the requirement to uphold regulatory standards while also supporting procedures that improve patient care. The LUMC Liver Transplant Team recently implemented the use of the FDA-approved OrganOx Metra normothermic machine perfusion instrument for maintaining donor livers under physiologic conditions prior to transplantation. The team mitigates risk for biliary complications associated with fibrin microthrombi by perfusing a combination of alteplase rTPA with fresh frozen plasma and monitoring the prevalence of fibrin through the release of D-dimers over the course of ex situ normothermic preservation. Liver perfusate is not an FDA-approved specimen type for the analysis of D-dimer and requires laboratory verification as a compatible specimen for analysis.

Methods

The D-dimer HS 500 assay was performed on an ACL TOP 750 analyzer. Carry-over studies evaluated activated partial thromboplastin time (aPTT) of a normal control (citrated plasma) specimen initially, and repeatedly in-between sets of D-dimer measurements of heparinized perfusate specimen. Recovery studies were performed by mixing liver perfusate with citrated plasma spiked with D-dimer in known ratios, allowing for the calculation of an expected value of D-dimer.

Results

Carry-over studies measuring aPTTs of normal plasma specimens provided %CVs of 1 and 3%. Recovery of D-dimer in seven sets of independent add-mixture series (three mixed specimen per set) was on average 105% with a range from 95 to 113% recovery.

Conclusions

Concern for cross-contamination of perfusate specimen containing heparin anticoagulant on analyzers used to evaluate routine coagulation testing was mitigated by the results of the carry-over study, which provided %CV within the manufacturer limit for intraday precision. Recovery studies evaluating matrix effect for the analysis of D-dimer on liver perfusate generated under the LUMC liver transplant protocol indicated that the matrix of the perfusate did not impact D-dimer analysis, as compared to the FDA-approved specimen type.
A-159 Re-evaluating Ferritin Cutoffs for the Diagnosis of Iron Deficiency and Iron Deficiency Anemia
Authors: Troike K , McShane A J
Published: October 2024

Background

Iron deficiency (ID) is the most common nutrient deficiency in the world and iron deficiency anemia (IDA) is estimated to affect 1.24 billion people. Serum ferritin is an iron storage protein commonly used as a biomarker for ID and IDA. The ferritin lower limit of normal (LLN) defined in many clinical laboratories is lower than evidence-based cutoffs and may not detect ID and IDA with adequate sensitivity, particularly when sex-based reference intervals (RIs) are used. This may be due in part to the use of asymptomatic, iron-depleted individuals for RI studies. Recent efforts have been made to increase the LLN for ferritin to at least 30 ng/mL and re-evaluate sex based RIs to improve diagnosis of ID and IDA. In this study, we aimed to investigate LLN ferritin cutoffs for ID and IDA in our patient population.

Methods

Patient data from June 2022-May 2023 were collected with approval from the Institutional Review Board (IRB), including age, sex, and values for ferritin, iron, transferrin saturation, and hemoglobin. Patients were classified as iron replete (IR), ID, or IDA based on measurements for iron, transferrin saturation, and hemoglobin. IR patients were defined as having iron and transferrin saturation values within current RIs, while those below the cutoff were grouped as ID. ID and IDA were further classified based on hemoglobin values. A range of ferritin concentrations between 1 and 100 ng/mL were used to calculate true and false positive rates (TPR and FPR) and diagnostic accuracy (DA). Receiver operator characteristic (ROC) curve and J index analysis were used to define optimal ferritin cutoffs for ID and IDA.

Results

Data from 26,455 patients (65% female) were used in this study.

Conclusions

These findings highlight the need to re-examine ferritin RIs in the context of ID/IDA, especially for those patients assigned female at birth.
A-161 Diagnosis of Lupus Anticoagulant Cases using Deep Neural Networks
Authors: Wang J , Leger R , Heikal N , Pruthi R , Chen D , Seheult J
Published: October 2024

Background

Laboratory identification of lupus anticoagulant (LAC) is critical for diagnosing antiphospholipid antibody syndrome, a common cause of unprovoked thromboembolic events and pregnancy morbidity. Commonly used clot-based assays for LAC include the dilute Russell’s viper venom time (DRVVT) and activated partial thromboplastin time (APTT), but these are susceptible to anticoagulant interferences. Despite standardization efforts, LAC interpretation remains challenging due to pre- and post-analytical complexity and limited expertise. This study explored the use of deep neural networks (DNN) to facilitate LAC diagnosis.

Methods

Retrospective data for LAC profiling with 13 features performed at Mayo Clinic between 2021 and 2022 was collected. The profiles of 7,202 patients were randomly divided into training (n=4608), validation (n=1153), and test cohorts (n=1441). Diagnostic outputs including LAC positivity (determined by DRVVT or APTT) and the presence of warfarin (WAR) and heparin (HEP), were confirmed by expert assessment. Three DNN architectures were developed and evaluated (Figure 1): (1) four single output DNN models with a domain knowledge driven selection of input features, (2) four single output DNN models using the same 13 inputs across all models, and (3) a multi-output DNN model. Model performance was evaluated using the F1 score and the index of balanced accuracy (IBA) for each output label.

Results

Architecture 1 achieved high diagnostic accuracy with IBA values of 0.989-1.000 for all four outputs and F1 scores of 0.945-0.991. Architecture 2 exhibited F1 scores of 0.902-0.973 and IBA values of 0.935-0.992. Architecture 3 achieved F1 scores of 0.933-0.967 and IBA values of 0.941-0.983.

Conclusions

All three architectures accurately classify LAC and common anticoagulation effects. The comparable performance of architectures 1 and 2 suggests that manual feature extraction is unnecessary. Given its relative simplicity and versatility, architecture 3 is the preferred choice for implementation in a clinical laboratory to standardize LAC diagnosis.
A-164 Establishment and Verification of Reference Intervals for Red Blood Cell Series and Total Leukocytes in Whole Blood Counts of Chilean Adults Using refineR and Multi Criteria Expectation Maximization Methods
Authors: Prieto C , Delgado H , Dias A C , Stevenson M , Díaz N , Arias S , Apablaza C
Published: October 2024

Background

This study aimed to define RIs for red blood cell series and total leukocytes in the complete blood count, in adult patients.

Methods

We used a sample of 822 patients aged 18 to 60 years, both female and male. The study focused on the red blood cell series (RBC Count, hemoglobin, Hematocrit, MCV, MCH, MCHC and RDW) and total leukocytes. RIs and respective 90% confidence intervals were estimated using the refineR method and a multi-criteria method combining the Expectation Maximization algorithm, mode, antimode analysis, an adapted Horn algorithm (5 cycles of Tukey's test with intermediate trimming and 2 cycles of Tukey's test with Box-Cox transformed data if applicable after analyzing Lambda, kurtosis, and skewness coefficients) and distribution truncation based on clusters and antimode. The RIs estimates using refineR and multi-criteria methods were compared to the manufacturer's specifications (Mindray). Flagging Rates were used to validate the method that estimates the most representative RI for the population served by the laboratory. Equivalence Interval and Bias Ratio were utilized to assess the practical relevance of sex-based partitioning.

Results

The method based on the multicriteria approach demonstrated the best performance, with flagging rates showing results lower than 4.7% and higher than 0.9% after the truncation of the results distribution. In general, the refineR method generated very narrow RIs, while the manufacturer's specifications resulted in very wide or shifted RIs, which do not adequately represent the data distribution profile. It was identified the need to partition by sex for the parameters Red Blood Cell Count, Hemoglobin, and Hematocrit, which is in agreement with the manufacturer's specifications.

Conclusions

The estimated RIs better represent the laboratory-served population, highlighting the importance of adhering to ISO 15189 normative requirements. This involves verifying and/or determining RIs to provide representative information of the served population, thereby aiding in medical decision-making.
A-179 Improving specimen labeling errors in the pediatric emergency department at a tertiary care hospital
Authors: Wang L , Huang J , Wu Y , Chow S , Fernado J , Kim C
Published: October 2024

Background

Specimen labeling errors, such as unlabeled, mislabeled, or sample-requisition mismatches, pose risks to patient care. Correcting these errors, particularly for pediatric patients, is crucial to avoid unnecessary stress to the family and time spent on follow-up. In 2018, the BCCH Emergency Department (ED) reported the highest labeling error rates among all departments at our tertiary care hospital, accounting for approximately 30% of all errors.

Methods

To address this issue, a quality improvement project following the Model for Improvement framework was initiated in 2019 at the ED. An interdisciplinary team approach was adopted initially, and a positive patient identification system (PPID) was implemented later in February 2022. Patient Safety Learning System (PSLS) data from January 2019 to November 2023 were analyzed to evaluate the impact of these interventions.

Results

1763 (22.6%) of the 7802 PSLS events recorded during this period were sample labeling errors. Ward collections accounted for the majority of labeling errors compared to laboratory collections (20.3% vs 2.4%). In the ED, most of the labeling errors occurred with unspecified samples (25%), swabs (24%), urine (23%), and blood culture (19%). The interdisciplinary team intervention initially reduced labeling errors, but its impact was inconsistent. However, the implementation of PPID in 2022 led to a significant and consistent decrease in labeling errors in the ED (4.51 errors/month vs 1.14 errors/month; p = 0.01) (Figure. 1).

Conclusions

While the implementation of PPID has notably improved labeling errors in the ED, complete elimination remains challenging. In addition, more specific PSLS filing categories are needed to capture sources of labeling error from unspecified samples. Continued efforts are necessary to achieve the goal of zero labeling errors.
A-189 Six Sigma for Thyroid Stimulating Hormone (TSH)
Authors: Ramadan-Boscolo D R , Utiyama A H , de Andrade S S , Tufik S
Published: October 2024

Background

Effective quality control processes within clinical laboratories are crucial for monitoring and assessing the analytical performance in terms of precision and accuracy. The regular measurement of these parameters plays a pivotal role in ensuring patient safety and is often a prerequisite for accreditation by various organizations. Six Sigma metrics have emerged as one of the most efficient tools for evaluating the analytical performance of clinical laboratory systems. This approach integrates Total Allowable Error (TEa), encompassing both random and systematic errors into a single comprehensive index. This study details the establishment of Six Sigma in our laboratory for Thyroid Stimulating Hormone (TSH) for eight different modules of Siemens Atellica IM.

Methods

A one-year dataset was compiled from the assessment of internal quality control for evaluating the imprecision (expressed as coefficient of variation - CV%) and Bio Rad’s Unity Interlaboratory Program for inaccuracy (quantified as bias%) in TSH test processed across eight modules of Atellica IM (Siemens). The data was categorized into two groups based on QC levels. The TEa utilized in this study was set at 20% according to CLIA standards. Six Sigma metrics were subsequently calculated for each QC level and equipment based on TEa, CV%, and bias%, utilizing the formula Sigma=(TEa - bias%)/CV%.

Results

Six Sigma values calculated for the QC level 1 ranged from 4.92 to 6.20 across all equipment, while for the QC level 2 Six Sigma varied from 4.87 to 7.59. All values found were classified as Good, Excellent and World Class according to the normalized method decision chart. These results showed that analytical performance of all eight modules of Atellica IM were similar in terms of random and systematic error.

Conclusions

Six Sigma serves as potent tool for monitoring the analytical performance of the TSH test. It is crucial to independently track different levels of QC in TSH testing for enhanced control over the analytical system. The performance of Siemens TSH test in our laboratory has proven to be very satisfactory.
A-194 Point of Care Glucose Test Time Interval Data Used as a Proxy Indicator for Glucose Meter Cleaning and Disinfection Adequacy Between Patient Testing Episodes
Authors: Reineks E Z , Carr M F
Published: October 2024

Background

At a community hospital within our health system (269 inpatient beds and 46 glucose meters), a condition level finding was cited for blood found on one meter during a routine regulatory survey. Point of Care (POC) test instrument portability is often an essential characteristic in choosing to perform a POC test at the bedside. Glucose meters are in close proximity to patients who are providing fingerstick blood samples for immediate testing. The process of obtaining the blood sample and applying it to the test strip may result in blood inadvertently getting on a caregiver’s gloves and the glucose meter itself. Per the testing protocol, when testing different patients consecutively, the caregiver safely discards the used test strip, changes gloves, and performs the cleaning and disinfecting (C&D) of the glucose meter to ensure the next patient is not exposed to blood from the previous test. The C&D procedure requires a minimum two minutes of drying time. We estimate that consecutive POC glucose tests should have about a three minute gap when the meter gets proper C&D between patients. We sought to determine whether the time interval between tests could be used an indicator of improved C&D compliance during a process improvement project.

Methods

As a patient safety concern, improved compliance with C&D steps was needed. Various stakeholder groups conducted a three month long campaign of education, training, and surveillance to improve C&D compliance. As a proxy for monitoring the C&D, data on test times from individual glucose meters was downloaded from the POC middleware system. Sample information, patient identifiers, and test results were not obtained. Time intervals between tests were calculated and the number of tests that occurred on the same instrument with short intervals (< 3minutes) were counted.

Results

Data from the POC middleware system was downloaded from the month prior to and immediately following the project (July and November). In July, there were 733 instances of glucose tests being carried out within three minutes of a previous test on the same instrument, from a total of 6622 tests (11.1%). In November, there were 565 occurrences (out of 6445 tests, or 8.8%), a decrease of 23% from the beginning count, and a drop of 2.3% among all tests. A follow up survey found no instances of blood on meters.

Conclusions

Using the time between tests interval as a proxy for monitoring glucose meter C&D is straightforward when the data is readily available. In our application of this process, an improvement in the number of short intervals was found, but the number of short intervals was not completely eliminated. This is likely due to occurrences when a test was repeated on the same patient or when training or competency was taking place, circumstances in which C&D not required. To observe patient privacy, our data collection approach could not eliminate these potential confounders of our data.
A-199 Use of R programming for monthly quality control (QC) data review in a university hospital laboratory
Authors: Stickle D , Toohey J M
Published: October 2024

Background

Our laboratory information system (LIS) provides only cursory capability for monthly review of quality control measurements. We streamlined the major portion of this process using customized programming in R, as described below.

Methods

Month-long QC data were obtained using the LIS archive’s data retrieval program (QLIK). Data were read and processed using our R program (QC-R.R), for each analyte and QC level, to produce both Levey-Jennings (LJ) plots (for evaluation of possible temporal trends in QC data) as well as cumulative distribution (CD) plots (to evaluate whether QC data are normally distributed). Plots were sent to graphics files whereby they could be reviewed sequentially by a single forward-arrow click.

Results

Our laboratory utilizes two QC measurements for each of 157 analytes. Thus, monthly review involves evaluation of 314 datasets for QC results. With the R program outputs, results can be reviewed visually in rapid fashion, to determine whether there exist discrepancies between fixed limits and current data. An example CD plot is shown in Figure. In this example, QC data (points) were, appropriately, normally distributed (theoretical curved line based on data mean and SD). The currently operative fixed mean±2SD limits (vertical dashed lines) were, however, disparate from those of data (vertical solid lines). This set of results was thus marked for updating of mean and SD, with reference to a master list of results of calculations generated by the QC-R.R program. Time for review of monthly QC results was dramatically reduced, from more than 12 man-hours to less than 4 man-hours per month.

Conclusions

Limitations of our LIS for monthly QC review were bypassed using R-based programming to automate a significant fraction of the process. This led to a significant savings in labor attending to this important task. The use of CD plots is helpful for rapid data interpretation.
A-206 Exploring the Surge in Clinical Laboratory Inflation: A Comprehensive Review
Authors: Jurinic E
Published: October 2024

Background

The healthcare industry has been experiencing significant inflationary trends, particularly in clinical laboratory services. This surge in inflation is attributed to various factors, including the increased demand for laboratory services, changes in the practice of clinical laboratory science due to technological advances, and vacancy rates that exceed the number of Medical Laboratory Science (MLS) graduates. Additionally, supply chain shortages, increased operational costs, and staffing budget increases, further exacerbating the inflationary pressures on clinical laboratory services.

Methods

The review of the factors behind the surge in clinical laboratory inflation involved a comprehensive review of industry data and insights from supply chain leaders, and recent surveys conducted by the author sent to 50 laboratory leaders with a response rate of 50%, Data from the Bureau of Labor Statistics (BLS) and industry interviews were integrated to provide an understanding of the inflationary trends affecting the healthcare industry, with a specific focus on clinical laboratory services.

Results

The review revealed that domestically produced healthcare inputs, labor, diagnostics, and lab equipment have seen dramatic price increases. For instance, prices manufacturers charge for diagnostic reagents for testing have increased by 9.2%, while lab equipment prices have risen by 7.1%. Some product lines have experienced increases of 15-20%. Moreover, the post pandemic has led to attrition rates climbing from 10%-37% to the current rate of 35%-61%, resulting in 30%-50% staffing budget increases. 36% of lab leaders feel Group Purchasing Organizations (GPO’s) are incentivized to allow increases, it’s estimated that as much as 80% of GPO revenue is generated by fees paid by vendors. 28% feel the vendors are passing along but not sharing in the inflationary pressures as reimbursements decline. 20% feel it’s an excuse for vendors to pad margins. 16% feel that it’s due to lack resources or lab specific procurement expertise in supply chain departments.

Conclusions

These factors have contributed to the significant surge in clinical laboratory inflation. The surge in is a complex phenomenon influenced by multiple interconnected factors. Addressing this inflationary trend will require a collaborative approach that considers the impact of supply chain shortages, increased operational costs, and staffing. Proactive measures to mitigate the adverse effects of inflation on clinical laboratory services, such as ensuring reliable data in negotiations, addressing the workforce shortage and strong negotiations with vendors will be essential to sustain the cost, quality, and accessibility of these critical healthcare services.
A-212 Comparison of formulas for measuring LDL in cancer patients with hypertriglyceridemia
Authors: Sánchez -Jacinto B , Arroyo- Huanaco O , Pizarro-Robles R
Published: October 2024

Background

Treatments in recent years in cancer patients have been very favorable. However, some data suggest that dyslipidemia plays a role in carcinogenesis through the deregulation of the lipid signal, and this possibly leads to the development of cardiovascular diseases. A marker for the diagnosis and treatment of dyslipidemias is low density lipoproteins (LDL), and currently, the primary care level still calculates LDL due to the high costs of the direct method; Friedewald has been used since 1972, and in recent years, new equations have appeared such as Sampson and Martin - Hopkins (MH). These formulas use total cholesterol (TC), high-density lipoproteins (HDL), and triglycerides (TG). Objective was to compare the formulas for measuring LDL in cancer patients with hypertriglyceridemia.

Methods

It´s an observational, cross-sectional - and analytical study. The sample size was 880 lipid profiles processed in an automated dry chemistry analyzer between January and March 2023 at an Oncology Institute in Lima, Peru. The population was divided into several groups of triglyceride levels (<200 mg/dl, 200- 399 mg/dl, ≥ 400 mg/dl), and LDL was stratified according to ATP III to evaluate the classification of patients with respect to LDL formulas. The statistical analysis was performed by descriptive analysis of the categorical and numerical variables; The correlation was performed using Pearson, and a comparison of the LDL formulas was used by Bland Altman to determine the bias and percentage bias. To clinical relevance, it was compared with the biological variation of the Westgard table. A p-value <0.05 was considered significant, and the data were processed in MedCalc v20.027.

Results

The average age of the patients was 55.02 ± 13.92 years, and the frequency of female sex was 77.39% (n = 681). The average measured TC, HDL, and LDL were 208.95 ± 54.84 mg/dl, 41.53 ± 13.83 mg/dl, and 115.70 ±45.53 mg/dl, respectively, while the calculated average LDL in Friedewald was 120.40 ± 48.57, MH was 130.49 ± 44.15 mg/dl and Sampson was 125.97 ± 45.53 mg/dl. The correlations of measured versus calculated LDL were strong (r > 0.89) at different TG levels; Statistical (p<0.05) and clinical relevance was observed in the Friedewald formula when TG ≥ 400 mg/dl, while the MH and Sampson formulas presented clinical and significant relevance at all triglyceride levels. Patients with an LDL level between 130 and 159 mg/dl presented a more substantial classification error according to the Friedewald, MH, and Sampson formulas.

Conclusions

Friedewald formula presented a clinically acceptable bias up to 400 mg/dl, unlike the MH and Sampson formulas, which were affected by triglycerides ≥ 150 mg/dl.
A-220 Method Intercomparison Study between Atellica Solution® and CI® Analyzers for HDL and LDL Methods
Authors: Ruiz-Alvarez M , Lapeña-Garcia S , Garcia-Valdecasas S , Costales-Lucia T , Sanchez-Fuertes J , Barrionuevo-Gonzalez M
Published: October 2024

Background

After validating a new Siemens equipment, the Atellica CI®, it is essential to ensure result interchangeability in our laboratory accredited by ISO 15189:2012.

Objectives

verify if the results of the HDL and LDL methods measured by Atellica Solution® and CI® analyzers are interchangeable and could be used interchangeably on both instruments

Methods

40 patient samples were selected for each assay, with results distributed across the entire measurement range and analyzed by both instruments. Result evaluation was conducted using the statistical program Method Validator v.19. To determine the degree of agreement between the instruments, a Bland-Altman mean difference analysis and a Passing-Bablok regression were performed. Statistics are expressed with their 95% confidence intervals (CI).

Results

HDL: After analyzing the Bland-Altman differences, a systematic error was found as the mean difference between the results obtained by Atellica Solution® and CI® was 0.686, with higher results in CI 1900®. Regarding the Passing Bablok analysis, constant systematic differences were observed as the confidence interval of the intercept does not include the zero value, and statistically significant proportional differences based on the slope whose 95% CI does not include the value 1.LDL: After analyzing the Bland-Altman differences, a systematic error was found as the mean difference between the results obtained by Atellica and CI® was -3.16, with higher results in Atellica Solution®. Regarding the Passing Bablok analysis, there were no systematic differences of constant type as the confidence interval of the intercept includes the zero value, but there were proportional differences based on the slope whose 95% CI does not include the value 1.

Conclusions

After evaluating the results, it is concluded that Atellica Solution® and CI® analyzers are interchangeable for HDL and LDL assays despite the presence of a systematic error, as the observed difference is not clinically significant.
A-226 Validation of Calculated Glycerol Corrected Triglycerides
Authors: Scott R , Meeusen J W , Jaffe A S , Vasile V C , Donato L J
Published: October 2024

Background

Patients with elevated free glycerol can have falsely elevated triglyceride concentrations known as pseudohypertriglyceridemia. Most clinical laboratory triglycerides methods measure the concentration of glycerol released following the enzymatic hydrolysis of triglycerides to quantitate triglycerides. Thus, increased free glycerol can lead to falsely increased triglyceride concentration. Identifying and measuring the interference is necessary to effectively manage patient care interventions. A glycerol-corrected triglyceride assay was discontinued from Roche Diagnostics in 2021. We created a process to determine glycerol-corrected triglycerides by measuring glycerol directly using an assay from Randox Laboratories.

Methods

Glycerol (Randox Laboratories) and nonglycerol-blanked triglycerides (Roche Diagnostics) were measured in serum using the Roche Cobas c501. Stability (n=10) and expected values (n=206) were determined using serum from healthy volunteers. Linearity (n=5) and precision (5 replicates on 2 runs over 5 days) were determined using residual clinical serum spiked with exogenous glycerol as necessary to span the analytical measuring range. Glycerol corrected triglyceride concentrations were calculated by subtracting glycerol (mmol/L) from triglyceride (mmol/L) and converting to mg/dL (x88.59). Samples previously measured using the Triglyceride/Glycerol Blanked Reagent (n=53, Roche Diagnostics) were used to assess accuracy. (Figure 1)

Results

Glycerol measures were stable 3 days refrigerated and 30 days frozen. Precision coefficient of variation was 1.0% within run and 7.1% within lab. Expected values were 10 - 109 mcmol/L, and the method was linear between 2 - 3,971 mcmol/L. The average bias between methods was 11.0% with slope of 1.02 and R2>0.97 (Figure 1).

Conclusions

Glycerol can be accurately measured using the Randox method and the result can be used to generate an accurate glycerol-corrected triglyceride concentration.
A-228 Evaluation of an Equation for Low-Density Lipoproteins-Triglycerides in Predicting ASCVD Risk in the UK Biobank Population
Authors: Wolska A , Sampson M , Zubiran R , Meeusen J W , Donato L J , Jaffe A S , Remaley A T
Published: October 2024

Background

Low-density lipoprotein-cholesterol (LDL-C) is the main univariate risk marker recommended for initial atherosclerotic cardiovascular disease (ASCVD) risk stratification. LDL, however, transports other constituents that can serve as pro-atherogenic risk markers. Like LDL-C, the triglyceride content of LDL (LDL-TG) can also be estimated from the lipid panel. Using data from UK Biobank (UKB), we compared LDL-C to LDL-TG for predicting future ASCVD events.

Methods

A UKB dataset (N=270,436) of primary prevention individuals not on lipid lowering therapy was used for analysis. LDL-C was calculated by the following equations: Friedewald, Extended Martin-Hopkins, Sampson-NIH, and Enhanced Sampson-NIH. LDL-TG was calculated by our newly devised equation (eLDL-TG = TG/38.46 + nonHDL-C/5.75 + (TG/nonHDL-C)/0.103 + (1/HDL-C)/0.0041 - 2.95)), which was derived from a dyslipidemic cohort (N=40,202) analyzed by the beta-quantification reference method.

Results

Although there was only a modest comparison between eLDL-TG and measured LDL-TG (R2=0.601, slope=0.611), eLDL-TG appeared to be a better risk marker than LDL-C in UKB. When the population was divided into quintiles and analyzed by survival curve analysis, eLDL-TG was better than LDL-C in segregating patients into low- and high-risk groups (Figure). Similar findings on the relative value of LDL-TG over other risk markers were also found when analyzed by ROC analysis (AUC: eLDL-TG>HDL-C>remnant-cholesterol>NonHDL-C>TG>apoB>Sampson-NIH-LDL-C). By Cox-proportional hazard analysis eLDL-TG showed alongside apoB and NonHDL-C the strongest association with future ASCVD events. Discordance analysis showed that when stratified as LDL-TG/LDL-C as High/Low there was an 26% increased risk for ASCVD, and when as High/High showed an 40% increased risk for ASCVD.

Conclusions

Like LDL-C, LDL-TG levels can also be estimated from the standard lipid panel. If our findings hold in other populations, the superiority of LDL-TG over LDL-C as a risk marker suggest that the reporting of calculated LDL-TG along with LDL-C could be a new strategy for improving ASCVD risk stratification.
A-233 Wastewater-based surveillance for SARS-CoV-2 Subvariants using the Nanopore Sequencing in South Korea
Authors: Kim Y , Noh T , Lee S , Kim Y , Lee K
Published: October 2024

Background

Genomic tools to identify and characterize SARS-CoV-2 and its various subvariants were adapted from clinical settings. Wastewater-based surveillance (WBS) has since been recognized as tool of transformative potential, providing real-time objective, comprehensive data on the prevalence and distribution of the target pathogen across the space and time. We conducted wastewater-based epidemiology (WBE) at sewage treatment plants to find a more efficient mutation monitoring method, and respond to pre-existing infectious diseases.

Methods

In our study, domestic sewage collected every weeks from 6 sewage treatment plants in urban region from June 2023 to November 2023 was used to concentrate and isolate viruses using wastewater ultra nucleic acid isolation kits with virus enrichment kits. Positive samples confirmed were subjected to SARS-CoV-2 whole genome sequencing (WGS) using GridION (Oxford Nanopore Technologies). The experimental data was aligned through Epi2me, analyzed using Nextclade, that was compared with KCDC official reports and our monitoring results (total 780 cases) ongoing at SCL.

Results

Various mutations were identified at each collection time, occurring new subvariants over time. The mutations (orf1a:A690V, S:F456L) sequentially confirmed from the sample collected on Aug. 23rd are specific mutations of XBB.1.9.2 and EG.5. In addition, the mutations orf1b: D54N, S:L455F, which were sequentially confirmed from the sample collected on Sep. 6th, are HK.3, a descendent lineage of EG.5, and FLip variants, which show growth advantage, were confirmed. Furthermore, the detection rate of FLip variants detected from Sep.6th gradually increased from 17% to 67% (Fig.1). Taken together, we confirmed the similarity between monthly detected mutations and changes in dominant species over time by comparing with ‘monthly COVID-19 mutation trends’ and ‘HK.3 dominance announcement by KCDC’(Fig.2).

Conclusions

WBS would be a useful tool for proactively responding to infectious diseases including COVID-19 by exploring new mutations and predicting changes in dominant species.
A-238 Lyophilized Bead Multiplex Assays for CT/NG/TV Detection
Authors: Moore R , Tang N , Tran E , Fisher T , Kadam R , Wang J , Farmer A
Published: October 2024

Background

Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), and Trichomonas vaginalis (TV) cause the three most prevalent sexually transmitted infectious diseases in developed countries, contributing to approximately 300 million new infections worldwide, annually. Due to the challenges of culturing and using immunoassays to detect these pathogens, DNA-based molecular tests have been instrumental, and current trends in molecular testing for CT/NG/TV have moved toward point-of-care tests.

Lyophilization provides benefits required for point-of-care tests, such as ambient storage, extended shelf life, simplified downstream workflow, and variable formats. A lyophilized-bead format enables an assay to be placed in sealed consumables and stored at room temperature for up to two years.

An additional consideration is the need for endogenous and exogenous controls. Exogenous in-process controls can be used to validate the performance of workflows culminating in a qPCR reaction and provide flexibility to test developers. An ideal exogenous in-process control is nonhomologous to human or human pathogen genome sequences and does not align with primers or probes designed for pathogen detection.

The objective of the current study was to develop a lyophilized-bead-format assay containing an ideal exogenous control and establish whether it is a valid option for the accurate detection of CT, NG, and TV.

Methods

A synthetic DNA control containing targets for CT/NG/TV/RNase P (RP)/exogenous control was used to establish an analytical LOD by qPCR. The 5-plex assay, lyophilized in beads, was hydrated and tested for linearity over an 8-log range of the synthetic DNA control. Further, the qPCR lyophilized bead with assay was tested with a proficiency sample by extracting DNA from ZeptoMetrix NATtrol CT/NG/TV Positive Control and spiking-in the exogenous DNA control. qPCR was performed on the extracted DNA and human cDNA.

Results

The analytical LOD for the control DNA was determined to be 5 copies/reaction. Additionally, linearity was observed by qPCR over an 8-log range of the synthetic DNA control for all five targets in multiplex with good efficiencies and R2 values. Finally, all targets were amplified and detected by qPCR using DNA extracted from ZeptoMetrix NATtrol CT/NG/TV Positive Control with spiked-in exogenous DNA control. The detection of the exogenous sequence in the qPCR multiplex reaction indicated the extraction was successful.

Conclusions

We have developed a multiplex qPCR assay in lyophilized-bead format that can identify infectious agents, such as CT/NG/TV, in a single reaction. We have included RP as a human host control, plus an exogenous in-process control, for a total of 5-plex in separate fluorescent channels. We have shown that our lyophilized beads with assay produce a positive qPCR result when combined with a proficiency sample.
A-240 Validation of the OSOM Flu SARS-CoV-2 Combo Home Test
Authors: Yaji S , Shimizu K , Fong D , Gonzalez E , Aihara T , Delgado Flores N , Dennis K , Nguyen-Brown T , Cox K , Baytan A , Schenken J , Morita M , Chun A , Nair-Schaef D
Published: October 2024

Background

COVID-19 and Influenza respiratory infections often cause similar symptoms, however, treatment is different for each infection. Delay in diagnosis can be fatal in some cases, therefore rapid detection and differentiation are critical for appropriate medical treatment. The OSOM Flu SARS-CoV-2 Combo Home Test is a lateral flow immunoassay intended for the qualitative and differential detection of nucleocapsid protein from SARS-CoV-2 and influenza A/B proteins using self-collected anterior nasal swab samples (self-collection is only recommended for those 14 years of age and older). An interpretation card, included with the test, is used to reliably distinguish among possible testing results.

Methods

A prospective study was conducted at multiple geographically distinct sites in the U.S during the 2023-2024 season for the validation of the OSOM Flu SARS-CoV-2 Combo Home Test to detect SARS-CoV-2/Flu A/Flu B in self-collected anterior nasal swab samples. The study evaluated the performance in symptomatic individuals only. A total of 703 prospective samples were evaluable for SARS. For Flu A/B, four additional subject samples were unevaluable due to comparator results not being available and a total of 699 samples were included in the data analysis. The comparator tests were FDA 510(k)-cleared molecular assays.

Results

Influenza A was detected with a sensitivity of 93.1% (95% CI: 84.8%-97.0%) and a specificity of 99.5% (95% CI: 98.6% - 99.8%). Influenza B was detected with a sensitivity of 89.1% (95% CI: 77.0% - 95.3%) and a specificity of 99.7% (95% CI: 98.9% - 99.9%). SARS CoV-2 was detected with a sensitivity of 87.0% (95% CI 78.6% - 92.4%) and a specificity of 99.1% (95% CI: 97.9% - 99.6%) when Ct values are standardized and 10% of samples are considered low positives, having a Ct value of ≥ 30.

Conclusions

With respiratory infections often showing indistinguishable symptoms and the possibility of co infection, lowering the testing hurdles for individuals to be able to perform a rapid differential test at home without the aid from a healthcare worker is the most effective way to mitigate spread and patient management from a public health perspective. Comparing the OSOM Flu SARS-CoV-2 Combo Home test to the FDA cleared comparators in a clinical setting, the OSOM Flu SARS-CoV-2 Combo Home test retains acceptable sensitivity and specificity. Using self-collected anterior nasal swabs which are minimally invasive alongside the low cost of a lateral flow immunochromatographic assay remains the most effective way to perform the serial screening testing recommended by the CDC in situations where transmission and severe outcome risk is high.
A-242 Assessing the efficacy of the one-step diagnostic algorithm for Hepatitis C
Authors: Bérgamo-Vázquez S , González-García L , González-Fernández C , Rives-Jimenez J , Prado-Ramos M , Guzmán-Olivares B , Enriquez-Marroquin C , Huerta-Velez A , Lao-López C , Álvarez-González M , Trapé-Pujol J
Published: October 2024

Background

Hepatitis C virus (HCV) infection can be asymptomatic and may go into spontaneous remission. However, in 60-80% of cases, it becomes chronic and progresses into cirrhosis or hepatocarcinoma. The virus is mainly transmitted parenterally, although cases of sexual and transplacental transmission have also been found. Since the development of direct-acting antiviral (DAA) therapy that can achieve a complete cure in 95% of patients, eradication strategies based on rapid diagnosis and treatment have been promoted. Therefore, the World Health Organization (WHO) has set a goal to eliminate HCV infection as public health threat by 2030. This is a retrospective study on the usefulness and effectiveness of the one-step HCV diagnostic algorithm application from May 2022 to December 2023 in our hospital.

Methods

The current HCV testing algorithm consists in a reflex HCV RNA testing, in which anti-HCV antibody-positive samples are automatically referred for HCV RNA testing on the same specimen. We used a serological electrochemiluminescence (ECLIA) assay for the detection of anti-HCV antibodies (Cobas e801, Roche Diagnostics®) in serum samples; and then a confirmation of HCV viremia by nucleic acid testing of HCV RNA in plasma samples (Cobas 4800, Roche Diagnostics®).

Results

A total of 22713 HCV serologies were performed with a total of 221 positive results (1%). Of the HCV cases, 21 patients had a newly diagnosis of active infection (10%) for whom we recommended derivation to the Digestive Service to initiate elimination treatment. 71% were men, with a mean age of 53 years and a standard deviation of 17. The mean age of women was 56 years with a standard deviation of 18. Coinfection with HIV or hepatitis B virus (HBV) was also assessed. Patients with HIV-HCV coinfection represented 9% of cases and were on average 12 years younger than HCV-monoinfected individuals. We did not identify any case of HBV-HCV co-infection.

Conclusions

The pattern we observed in terms of incidence by age and sex is similar to that reported in the literature, as well as the incidence of new HCV cases. One-step diagnosis enables the laboratory to identify the need for referring to a specialist, reducing the time to treatment for new and known but untreated cases. Therefore, global access to HCV diagnostics will be a keystone to success.