bluuebunny/crossref_metadata_2025_split huggingface dataset:
B-156 Impact of Seven Incremental Scenarios of QC Strategies
Authors: Brooks Z C 
Published: September 2023

Abstract

Background
CLSI EP23A states that “At the least, the ability of the QC procedures to detect medically allowable error should be evaluated.” It also defines that “The QC strategy using QC samples should include the following for each measuring system:

Methods
Eighty QC sample mean and SD values were compared to Peer Means and TEa limits for 14 routine chemistry analytes and four cardiac markers tested in December 2022. The number of patient results was normalized to 200 and 60 per day for routine and cardiac; the number reported before the first QC flag after a simulated shift to a 5% error was calculated using CatalystQC software for seven incremental scenarios of quality control beginning with [A] Original QC rules (1-3s/2-2s), chart limits, frequency (once per day), analytical bias & SD; [B] Westgard Rules based on sigma values; [C] Westgard QC frequency; [D] Chart means & SDs set to measured values; [E] Improved bias and/or SD where advised by CatalystQC software; [F] Single CatalystQC™ rules; [G] QC bracketed at the end of each day.

Results
The number of patients at risk decreased with each component of the QC process. Observed and advised number of QC samples per year was inconsistent.

Conclusion
Selecting Westgard rules and frequency alone will not reduce patient risk significantly. Assigning QC chart values to recent mean and SD and improving analytical bias and SD have the most impact on reduction of patient risk. Catalyst QC single rules are more effective than Westgard rules. Bracketing QC with verified-effective QC will reduce patient risk to zero.
B-157 Metrics to Express Risk in Terms of A. Risk Sources, B. Potential Events, C. Their Consequences and D. Their Likelihood.- As Recommended by ISO 31000
Authors: Brooks Z 
Published: September 2023

Abstract
Risk Metrics to express risk in terms of A. risk sources, B. potential events, C. their consequences and D. their likelihood (as recommended by ISO 31000) compared to sigma metrics.

Background
ISO 31000 Risk Management Guidelines provides guidelines on managing risk faced by all organizations. “Risk is usually expressed in terms of A. risk sources, B. potential events, C. their consequences, and D. their likelihood.” I compared risk metrics to implement these guidelines in medical laboratories to sigma metrics to examine the most effective way to enable risk evaluation as “the process of comparing the estimated risk against given risk criteria to determine the acceptability of the risk” (CLSO EP 23A).

Method
Eighty QC sample mean and SD values were compared to Peer Means and CLIA TEa limits for 14 routine chemistry analytes and four cardiac markers tested on two Ortho Vitros 5600 analyzers in December 2022. Risk evaluation was divided into Current Risk, driven by analytical accuracy and precision, and Potential Risk driven by effectiveness of the daily QC system to detect a simulated shift to a 5% error rate.
Current risk consequences were quantified as the number of failures per million opportunities and per year, and the estimated healthcare cost of lab error based on the costing process established for calcium in 2004.i
Potential risk consequences were quantified as the number of failures reported and healthcare cost of lab error following a simulated shift to create a 5% error rate.

Results

Conclusion
There is a good relationship between sigma and projected failures per year. Sigma is NOT a good indicator of cost of error per year, or the number or cost of failures reported after a simulated failure.
Endnotesi National Institute of Standards and Technology. Planning Report 04.1. The impact of calibration error in medical decision making.
B-158 Application of Statistical Analysis as a Tool to Monitor the Variation of SARS-CoV-2 and Other Respiratory Viruses
Authors: Carvalho G C , Nurmberger J M , Perez B B , Ramadan D R , Tufik S 
Published: September 2023

Abstract

Background
After the emergence of SARS-CoV-2 and its worldwide spread, only a few studies have correlated its co-circulation with other respiratory viral pathogens (RVP). The current report aims to identify the pattern of SARS-CoV-2 positivity in individuals co-infected with influenza A (FluA), influenza B (FluB), or respiratory syncytial virus (RSV) in São Paulo, Brazil.

Methods
Study Population and Characteristics: This study was a retrospective analysis from a clinical laboratory database in São Paulo, Brazil from January 15 to March 10, 2022. Test results for respiratory viral co-infections were recorded for individuals with SARS-CoV-2, FluA, FluB, or RSV. Nasopharyngeal swab samples were evaluated for the molecular diagnosis of respiratory virus infections using a real-time multiplex-PCR panel to search for specific genes for the detection of SARS-CoV-2, FluA, FluB, and RSV. Only samples positive for one or more pathogens (including SARS-CoV-2) were included in the study. We further stratified the specimens by SARS-CoV-2 positivity, comparing those that also tested positive for an RVP in each group. Statistical Analysis: Kruskal-Wallis analysis of variance followed by Dunn’s multiple comparison tests was used. Results with P &lt; 0.05 were considered statistically significant. Georeferencing analysis of SARS-CoV-2 mono-infection or co-infection with another respiratory virus was performed using an online available software application.

Results
A total of 28 932 samples were positive for at least one RVP, meeting the study criteria. Among those, a total of 93.13% were from individuals diagnosed with SARS-CoV-2 only, 2.99% positive for SARS-CoV-2 and FluA, and 0.99% positive for SARS-CoV-2 and RSV. There was no significant difference observed between genders nor individuals with mono-infection or co-infection. The analysis of age group showed a significant variation in positive cases between SARS-CoV-2 vs SARS-CoV-2 and FluA in the 0 to 10-year-old population (P = 0.017) and between SARS-CoV-2 vs SARS-CoV-2 and RSV in the 0 to 5-year-old population (P = 0.018). The highest number of positive samples for both SARS-CoV-2 and RSV was detected in the southern region of São Paulo (55.7%), compared to the north and east regions (44.3%).

Conclusion
Differences in diagnostic patterns of COVID-19 compared to other respiratory viral infections RVP co-infections are important for a better understanding of clinical outcomes, improved patient management, and better public health planning.
B-162 Direct Quantitation of Five Immunosuppressant Drugs in Dried Whole Blood Spots by a Fully Automated DSM-LC-MS System
Authors: Gibson R , Guo J , Samra S 
Published: September 2023

Abstract

Background
Liquid chromatography-mass spectrometry (LC-MS) is increasingly used in clinical research to quantify immunosuppressant drugs in whole blood as it can offer higher sensitivity and selectivity than other analytical techniques. LC-MS systems may be coupled with upstream dried matrix spot modules within one high-throughput, integrated and online workflow, allowing the extraction of matrices such as whole blood from pre-collected sample cards collected in a quick and minimally invasive manner.
In this poster, such a workflow will be utilized to demonstrate the quantitation of five commonly monitored therapeutic immunosuppressants; cyclosporin A, everolimus, mycophenolic acid, sirolimus and tacrolimus.

Methods
Calibration and QC samples were prepared in whole blood. 10 μL of each calibrant was then spotted in triplicate on DBS cards. Immunosuppressants and standards were extracted using a Thermo Scientifc™ Transcend™ DSX-1 Systems via flow-through desorption with a heated clamp. TurboFlow allowed interference removal in the extracted samples prior to analytical separation. An integrated software, Aria MX, controlled each step of sample desorption and separation. Analyte quantitation was performed by a TSQ Altis™ mass spectrometer, and the data was analyzed using TraceFinder™ 5.1 General Quan software.

Results
Five commonly therapeutically monitored immunosuppressant drugs were quantified from blood spotted onto DBS cards using a fully automated workflow, resulting in a highly sensitive 5-minute method. Calibration curves were fitted using a weighting factor of 1/x with requirements of R2 &gt; 0.98, RSD/CV &lt; 15%. The calibrants were used to determine the limits of detection (LODs) and quantitation (LOQs) shown in the below table.

Conclusion
A complete, integrated and fully automated workflow has been developed to rapidly and robustly quantify immunosuppressant drugs from dried blood spots. Quick and reproducible extraction of each target analyte and efficient chromatographic separation facilitated a 5-minute method for the quantitation of five of the most commonly therapeutically monitored immunosuppressant drugs.
B-171 Comparison of Intact PTH and PTH Assays using LC-MS/MS Among Non-Dialysis Dependent Chronic Kidney Disease Patients
Authors: Park H , Lee M , Lee K 
Published: September 2023

Abstract

Background
Parathyroid hormone (PTH) is an important regulator of calcium and phosphate homeostasis and bone remodeling. It is metabolized into PTH fragments, which are measured to a different extent by PTH assays of different generations because of differences in fragments recognized and lack of assay standardization. Therefore, we developed an LC-MS/MS method for 1-84 PTH and aimed to compare with immunoassay among patients with chronic kidney disease (CKD) stages 3 to 5 not yet on dialysis.

Methods
Blood samples were collected from 252 patients with CKD stages 3 to 5. PTH concentrations were measured simultaneously by using the second-generation—PTH intact and immunocapture in situ digestion LC-MS/MS spectrometry assays. The severity of the kidney disease was based on the eGFR calculated by using the Modification of Diet in Renal Disease (MDRD) formula.

Results
Serum 1-84 PTH concentrations were strongly correlated but significantly lower than the intact PTH concentrations (r = 0.956, P &lt; 0.0001). This finding was consistent among CKD stages 3 to 5. PTH concentrations by both assays (intact and 1-84 PTH) positively correlated with BUN (r = 0.613, r = 0.61; P &lt; 0.0001, respectively), phosphorus (r = 0.423, r = 0.427; P &lt; 0.0001, respectively) and negatively correlated with blood calcium (r = −0.355, r = −0.421; P &lt; 0.0001, respectively), protein (r = −0.289, r = −0.305; respectively, P &lt; 0.0001) and the estimated glomerular filtration rate (r = −0.468, r = −0.485; P &lt; 0.0001, respectively).

Conclusion
Among patients with CKD stages 3 to 5 not on dialysis, the 1-84 PTH assay detected significantly lower PTH concentrations compared with intact PTH assay. This LC-MS/MS method can provide accurate and precise PTH results in patients with severe chronic renal failure. Additional studies that correlate the diagnosis and management of CKD mineral and bone disorders are needed to determine whether 1-84 PTH assay results are better surrogate markers in these early stages of CKD.
B-174 Simple and Accurate Quantification of Four Direct Oral Anticoagulants in Plasma Using UPLC-MS/MS
Authors: Lin C , Huang Y , Cheng C , Yeh Y , Chang S 
Published: September 2023

Abstract

Background
Direct oral anticoagulants (DOACs) constitute first-line therapy used for many thromboembolic indications, such as prevention and treatment of venous thromboembolism and stroke prevention in atrial fibrillation. This class of medication consists of the direct thrombin inhibitor (dabigatran) or direct factor Xa inhibitors (apixaban, rivaroxaban, and edoxaban). DOACs quantification may be useful in the case of critical clinical situations, including drug accumulation in long-term treatment, overdosage, thrombotic or bleeding events. This study aimed to develop and validate an ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for the quantification of four DOACs: dabigatran, apixaban, rivaroxaban, and edoxaban in plasma.

Methods
The calibration curve samples were prepared by spiking drug free plasma with DOACs. The stable isotope labeled DOACs was used as an internal standard. After addition of the internal standard and protein precipitation, the supernatant was 10-fold diluted and injected into a chromatography system consisting of a UPLC BEH C18 (2.1 × 50 mm; particle size 1.7 µm) analytical column with gradient made of mobile phase (5 mM ammonium formate pH3.5 in water and methanol). The outlet of the column was connected to a triple quadrupole mass spectrometer with electrospray interface. Ions were detected in the positive multiple reaction monitoring mode. The concentration of analyte was calculated from the calibration curve and ion ratios between the analyte and the internal standard.

Results
The analytical range was linear with a correlation coefficient of over 0.99 in the range of 0–500 ng/mL for each analyte. Intra- and inter-assay imprecision were less than 3.1% (n = 20) and 6.0% (n = 20), respectively. The accuracy was evaluated by spike recovery and the mean recoveries were between 97.2% and 102.1% for all analytes. This assay showed no ion suppression or enhancement and no carryover. The chromatography run time was 4.5 min. The assay was applied to quantitate DOACs in plasma samples from 93 patients. Our data showed that the DOACs concentrations vary markedly individual patients after equal dose, this therapeutic drug monitoring of DOACs is essential to optimized drug efficacy.

Conclusion
An accurate, and cost-effective UPLC-MS/MS method for the quantification of DOACs was successfully applied for therapeutic drug monitoring in patients.
B-190 Development of an LC-MS/MS Method for Analysis of 7α-hydroxy-4-cholesten-3-one and Investigation Into its Utility as a Biomarker for Bile Acid Diarrhoea in the United Kingdom
Authors: Starbrook L , Kumar A , Ford C , Brookes M , Gama R 
Published: September 2023

Abstract

Background
Bile Acid Diarrhoea (BAD) is a common gastrointestinal condition estimated to affect 1% of the population. It is, however, significantly under-diagnosed. This is partly due to lack of diagnostic methods. In the United Kingdom (UK) the 75-SeHCAT retention scan is considered the gold-standard diagnostic method and is the only routinely available diagnostic test for BAD. SeHCAT retention of &lt;15% is considered consistent with BAD. The SeHCAT test is, however, expensive, time consuming, and exposes the patient to a dose of radiation. 7-alpha-hydroxy-4-cholesten-3-one (7αC4) is a precursor in the bile acid synthesis pathway and has demonstrated utility as a marker of BAD. There is limited data comparing 7αC4, which is measured in the United States, to the SeHCAT test, as SeHCAT is not licensed in the United States. 7αC4 can be measured by LC-MS/MS, thus can provide a more convenient, low cost and high throughput option for diagnosis of BAD.

Aims
1) To develop and validate an LC-MS/MS method for analysis of 7αC4. 2) To analyse serum samples taken from patients referred for SeHCAT scans. 3) To determine the utility of 7αC4 compared to SeHCAT for diagnosis of BAD.

Methods
The method was adapted from Donato et al. 750 µL of ice-cold acetonitrile containing 50 nmol/L D7-7αC4 as internal standard was added to 250 µL serum. Samples were vortexed for 30 s, centrifuged at 3000g for 10 min, and supernatant transferred into a vial for LC-MS/MS analysis. The LC-MS/MS system utilised a Gemini 5um NX-C18 column coupled to a Sciex 6500 TripleQuad mass spectrometer. Ethical approval was obtained to collect serum on patients undergoing SeHCAT retention scans for investigation of BAD. 40 patients who attended for SeHCAT scans had serum collected and stored at −80 degrees Celsius until analysis for 7αC4. Sensitivity and specificity of 7αC4 was calculated and ROC curve analysis performed.

Results
The 7αC4 method demonstrated an LoQ of 1.5 nmol/L, linearity up to 1095 nmol/L, and intra-assay %CV of 3.0%–7.7%. Using SeHCAT retention of &lt;15% as positive for BAD, ROC curve analysis demonstrated an AUC of 0.765. 7aC4 at a concentration of 175 nmol/L provided a sensitivity of 30% and a specificity of 92%. 7αC4 at a concentration of 50 nmol/L provided a sensitivity of 93% and a specificity of 46%.

Conclusion
The 7αC4 method demonstrated good analytical performance and was suitable for use in a routine laboratory. 7αC4 results of &lt;50 nmol/L suggest BAD is unlikely, and results of &gt;175 nmol/L make a diagnosis of BAD likely. Results in between these two values may be considered indeterminate and referral for SeHCAT scanning may be considered appropriate. The 7αC4 assay provides an opportunity to stratify patients for SeHCAT testing in the UK, improving diagnosis and management of BAD in patients with chronic diarrhoea and streamlining the patient pathway.
B-191 Development of a Simple, Robust, and Accurate LC-MS/MS Method for the Quantification of 17 Therapeutic Drugs: Initial Validation Results
Authors: Payto D A , Tang X , Giles R , Wei R , Colón-Franco J M 
Published: September 2023

Abstract

Background
Therapeutic Drug Monitoring (TDM) of Zonisamide (ZON), Felbamate (FELB), Lamotrigine (LMTR), 10-Hydroxycarbamazipine (OXCARB), Carbamazepine Epoxide (CARBEP), and Pentobarbital (PENTO) is essential in developing the regimen to produce the best clinical outcomes while monitoring for possible toxicity. Historically, both HPLC-UV and LC-MS/MS have been used for the quantification of these types of drugs. HPLC-UV is often subject to interferences and extended run times. Because of this, there has been a shift to LC-MS/MS methods which are often simple, fast, robust, and suffer from less interferences. Our objective was to develop a simple, robust, and accurate LC-MS/MS method to replace the HPLC-UV methodology.

Method
Serum (50 µL) and IS solution (100 µL; Zonisamide-13C6, Lamotrigine-13C,15N4, (±)-10,11-Dihydro-10-hychoxycarbamazepine-13C6, Carbamazepine-10,11-epoxide-13C6, Felbamate-D4, Pentobarbital-D5 in methanol with 0.5% acetic acid) were vortex mixed and centrifuged. Supernatant (50 µL) was mixed with 950 µL of water and vortexed mixed. Extract was injected on a Thermo Accucore C18 column. A quantifier and qualifier transition was monitored for all analytes. Multiple Reaction Monitoring (MRM) was performed in positive mode for all analytes except for PENTO (negative). Total chromatographic time for the 17 analytes is 7.31 min.

Results
No ion suppression, differential matrix effect, or interferences were observed. Analytical Measurement Range (AMR) data for the initial 6 analytes is presented in Table 1. The coefficient of variation (%CV) was &lt;2.8% for ZON, &lt;4.3% for FELB, &lt;7.3% for LMTR, &lt;3.8% for OXCARB, &lt;3.0% for CARBEP, and &lt;3.5% for PENTO. Method comparison with a previously validated method were acceptable (Table 1).

Conclusion
This simple, robust, and accurate LC-MS/MS method has been validated for the initial 6 anlytes and has been determined to be acceptable for clinical use. Validation of the remaining 11 analytes is ongoing (Pregabalin, Gabapentin, Levetiracetam, Ethosuximide, Lacosamide, Rufinamide, Primidone, Lamotrigine, Topiramate, Brivaracetam, and Perampanel).
B-201 Development of a Quantitative Gas Chromatography Tandem Mass Spectrometry Method for Metabolic Profiling of Urine Organic Acids
Authors: Ptolemy A S , Young K , MacDonald C , Garrett R , Kellogg M , Peake R W 
Published: September 2023

Abstract

Background
Urine organic acids (UOA) analysis is an essential investigation for inborn errors of metabolism (IEMs). The vast majority of methods use gas chromatography mass spectrometry (GC-MS) and are only qualitative. Gas chromatography tandem mass spectrometry (GC-MS/MS) offers greater sensitivity and selectivity, and enables the combination of qualitative screening with quantitative analysis. We describe the development and preliminary validation results of a quantitative GC-MS/MS method.

Methods
Urine creatinine concentrations were measured (CobasPro, Roche Diagnostics) and normalized to 1.25 mmol/L. After internal standard addition, each urine specimen, calibration standard and QC were incubated with a hydroxylamine solution followed by a liquid-liquid extraction. Further sample concentration permitted the isolated UOA to be derivatized using N,O-bis(trimethylsilyl)trifluoroacetamine (BSTFA) with 1% chlorotrimethylsilane (TMCS) prior to injection. GC-MS/MS analysis was performed using a GC-MS-TQ8050 triple quadrupole system (Shimadzu) with a 30 m DB-5MS (Agilent J&W) capillary column (0.25 mm, ID of 0.25 µm). Split injection mode was deployed with an initial column temperature of 70°C. The mass spectrometer was operated in electron ionization (EI) and multiple reaction monitoring (MRM) modes. The linear concentration ranges of the targeted metabolites within the devised GC-MS/MS protocol were determined.

Results
The optimal temperature gradient for high-resolution UOA separations was derived and their respective retention times determined. MS/MS detection parameters were optimized to permit the identification of precursor and production ions for all (N = 38) derivatized UOA. Total analysis time was 33.8 min per injection. The achieved linear concentration ranges for each metabolite was respectively determined. Linear concentration ranges and correlation coefficients of representative UOA: 3-hydroxyisovaleric acid (range = 9 to 708 µmol/L, R2 = 0.9874); lactic acid (range = 15 to 751 µmol/L, R2 = 0.9936); 3-hydroxybutyric acid (range = 15 to 751 µmol/L, R2 = 0.9991); and 2-methyl-3-hydroxybutyric acid (range = 10 to 501 µmol/L, R2 = 0.9996). The correlation coefficients for all remaining UOA ranged from R2 = 0.9458 (4-hydroxyphenylpyruvic acid) to R2 = 0.9998 (3-methylglutanoic acid and ethylmalonic acid).

Conclusion
The devised GC-MS/MS protocol permits the unbiased identification and quantification of UOA used to investigate IEMs. The linear concentration ranges for all analytes were determined and are appropriate for monitoring these markers of inherited enzyme deficiencies. The analytical performance of this UOA GC-MS/MS method must be further validated and its diagnostic performance for selected IEMs characterized.
B-213 Colorimetric LAMP as an Effective Tool for Rapid Molecular Diagnostics
Authors: Tanner N A , Li Z , Bruce J , Moncion R A , Cunningham C V , Langhorst B W , Carlow C K 
Published: September 2023

Abstract

Background
LAMP has become a standard molecular diagnostic technique, and its robustness and flexibility have enabled rapid, simple tests outside of traditional laboratory settings. The need for additional diagnostic methods became urgent during the SARS-CoV-2 pandemic, and LAMP was extensively evaluated and validated for numerous testing applications. We have extended the utility of LAMP to enable colorimetric detection, multiplex assays, variant identification, and robust direct tests for point of-care and workplace surveillance testing.

Methods
We utilized fluorescent and colorimetric RT-LAMP for assay development and our workplace screening program. A simple lysis buffer was applied to whole saliva samples and added directly to our colorimetric LAMP SARS-CoV-2 tests, with more than 100 000 samples tested. For variant calling we developed novel hybridization probes with single-base sensitivity to regions of sequence changes, and we characterized more than 5000 LAMP assays internally positioning mismatches in the LAMP primer regions to ensure robustness to and tolerance of mutations.

Results
The colorimetric LAMP assay displayed a sensitivity of 97% and specificity of 100%, and during its utilization in our in-house CLIA-certified workplace screening has identified &gt;300 unique positive cases derived from all variants of concern. 3654 blinded samples were analyzed by 3 operators and colorimetric readout was found to have a strong concordance with real-time fluorescence measurement (kappa = 0.999).

Conclusion
RT-LAMP is remarkably tolerant of sequence variation, with no individual positional mutation significantly affecting assay performance. Simple visual readout can be utilized for a reliable diagnostic indicator when sophisticated instrumentation is not available or desired.