bluuebunny/crossref_metadata_2025_split huggingface dataset:
B-031 Digital Elisatotal 25-Hydroxyvitamin D
Authors: Chae S , Chung J , Huh H , Moon S , Seo Y 
Published: September 2023

Abstract
Evaluation of digital ELISA using a high-resolution computational optical system for measurement of total 25-hydroxyvitamin D

Background
Biomarkers such as vitamin D occur at very low concentrations in blood, and sensitive detection methods are required in order to measure them. It is difficult to accurately measure low-concentration biomarkers using conventional enzyme-linked immunosorbent assays (ELISA) with fluorescence staining. By developing a digital ELISA (DELISA, Small Machines, Seoul, Korea) platform, a low concentration substance was realized by analyzing the digitalized signal through a high-resolution computational optical system in a microfluidic chip. We evaluated DELISA for measurement of total 25-hydroxyvitamin D (25OHD) in a fluidic chip.

Methods
The sample with monoclonal antibody bead conjugates mixture was incubated for 20 min at 37°C, then 30 uL sample mixture was applied to inlet of the chip. After additional incubation for 2 min at room temperature and 2 min after sample application, 120 uL of washing buffer was applied onto the washing hole and the chip was incubated for 10 min. 25OHD molecules in the sample bond to magnetic beads coupled with anti-25OHD at the end of the membrane. Chips were inserted into DELISA, and 25OHD results were derived by analysis of the computational method of microscopic imaging, proportional to the concentration of 25OHD. Assay performance characteristics of the limit of detection (LoD), precision and analytical measurement interval (AMI) using Vitamin D Depleted, Low and High Serum (Molecular Depot, San Diego, CA) were determined according to the Clinical and Laboratory Standards Institute (CLSI) guidelines. 87 Samples for method comparison studies were obtained from routine clinical samples submitted for 2 OHD determination or from apparently healthy normal volunteers. The study was approved by the Institutional Review Board of Dongguk University Ilsan Hospital (202207024).

Results
LoD was 10.3 ng/mL and within-laboratory precision was 11, 12, and 10% coefficient of variation at 14.1, 56.6, and 113.2 ng/mL, respectively. The best fit polynomial was a cubic equation from 14.1 to 113.2 ng/mL and the difference between the third-order equation and the first-order equation at Level 3 and 4 concentrations was outside the non-linearity limit. Comparison with chemiluminescent assay (Elecsys Vitamin D total II, Roche Diagnostics, Mannheim, Germany) yielded a 0.904 of correlation coefficient and 0.921 slope in the range from 7.0 to 57.9 ng/mL.

Conclusions
The DELISA 25OHD assay has a suitable LoD and precision, however, it is necessary to improve the AMI at high concentration values and correlation with the automated analyzer.
B-071 Comparison of Humoral and Cellular Immune Responses Between ChAd-BNT Heterologous Vaccination and BNT-BNT Homologous Vaccination Following the third BNT Dose: A Prospective Cohort Study
Authors: Kang H , Sim W , Jung J , Lee J , Ko G , Park H , Cha K , Choi J , Park K , Oh E 
Published: September 2023

Abstract

Background
The differential immune responses after two additional BNT162b2 (BNT) doses between ChAdOx1 nCoV-10 (ChAd)-primed and BNT-primed groups have not been elucidated. The aim of this study was to compare vaccine-induced humoral and cellular immune responses and evaluate breakthrough infection between the two vaccination strategies.

Methods
In 221 healthy subjects (111 in the ChAd group), longitudinal immune responses were monitored at 3, 4, and 6 months after the 2nd dose and 1, 3, and 6 months after the 3rd dose. Humoral immunity was measured by two fully automated chemiluminescent immunoassays (Elecsys and Abbott) and a surrogate virus neutralization test (sVNT). Cellular immunity was assessed by two interferon-γ (IFN-γ) release assays (QuantiFERON SARS-CoV-2 and Covi-FERON).

Results
After the 2nd dose of BNT vaccination, total antibody levels were higher in the ChAd group, but IgG antibody and sVNT results were higher in the BNT group. Following the 3rd dose vaccination, binding antibody titers were significantly elevated in both groups (ChAD-BNT; 15.4 to 17.8-fold, BNT-BNT; 22.2 to 24.6-fold), and the neutralizing capacity was increased by 1.3-fold in both cohorts. The ChAd-BNT group had lower omicron neutralization positivity than the BNT-BNT group (P = 0.001) at 6 months after the 3rd dose. Cellular responses to the spike antigen also showed 1.7 to 3.0-fold increases after the 3rd dose, which gradually declined to the levels equivalent to before the 3rd vaccination. The ChAd cohort tended to have higher IFN-γ level than the BNT cohort for 3–6 months after the 2nd and 3rd doses. The frequency of breakthrough infection was higher in the ChAd group (44.8%) than in the BNT group (28.1%) (P = 0.0219). Breakthrough infection induced increased humoral responses in both groups, and increase of cellular response was significant in the ChAd group.

Conclusion
Our study showed differential humoral and cellular immune responses between ChAd-BNT-BNT heterologous and BNT-BNT-BNT homologous vaccination cohorts. The occurrence of low antibody levels in the ChAd-primed cohort in the humoral immune response may be associated with an increased incidence of breakthrough infections. Further studies are needed on the benefits of enhanced cellular immunity in ChAd-primed cohorts.
B-111 Parallel Detection of Antibodies against CDR2L and CDR2 Increases the Diagnostic Specificity in Serological Diagnostics of Paraneoplastic Cerebellar Degeneration
Authors: Venkataraman I , Ott A , Scharf M , Miske R , Jäger D , Honnorat J , Komorowski L , Scheper T , Meyer W 
Published: September 2023

Abstract

Background
Onconeuronal anti-Yo antibodies are key diagnostic markers for paraneoplastic cerebellar degeneration (PCD) and often associated with gynaecologic/breast cancer. Cerebellar degeneration-related protein 2-like (CDR2L), but not cerebellar degeneration-related protein 2 (CDR2), has recently been shown to be the major anti-Yo target (1, 2). This study investigated if inclusion of CDR2L in diagnostic tests results in increased specificity for PCD.

Methods
Three panels were examined for reactivity against CDR2L and CDR2 by line blot analysis (EUROLINE). Panel A comprised 14 preselected sera from women with Yo-specific reactivity by indirect immunofluorescence assay (IFA) on rat cerebellum, presenting with cerebellar ataxia (n = 13) or peripheral neuropathy (n = 1) and underlying gynaecologic/breast cancers (n = 14). Panel B included 2132 non-selected consecutive serum/CSF samples from patients with neurological symptoms. Panel C consisted of 150 samples from healthy blood donors.

Results
In Panel A, all samples (100%) from patients with typical clinical characteristics for Yo-associated PCD showed strong positive reactivity to both CDR2L and CDR2. In Panel B, seven (0.3%) sera were anti-CDR2 negative, but anti-CDR2L positive, all of them from patients (6M/1F) without PCD and two with cancer or pre-cancerous condition. Eleven (0.5%) patients (9M/2F) were found to be anti-CDR2 positive, but anti-CDR2L negative, including two with cerebellar ataxia and three with cancer or pre-cancerous condition. In all 18 cases, IFA on rat cerebellum was negative, suggesting non-specific line blot reactivity to CDR2L or CDR2. In Panel C, 0/150 sera were anti-CDR2L and 1/150 anti-CDR2 positive, corresponding to specificities of 100% and 99.3%, respectively. The anti-CDR2 positive blood donor was male with negative IFA result.

Conclusion
Using CDR2L in addition to CDR2 increases the specificity in diagnosing anti-Yo-associated PCD. Positive line blot reactivity against both parameters seems to confirm the diagnosis. In contrast, results showing isolated reactivity against either CDR2L or CDR2 have to be interpreted carefully as they may apply to non-PCD cases.
B-131 The Prevalence and Clinical Relevance of the DFS Immunofluorescent Staining Pattern in a Large ANA-positive Cohort
Authors: Zhang Y , Rao L V 
Published: September 2023

Abstract

Background
The dense fine-speckled (DFS) nuclear pattern is one of the most common indirect immunofluorescence (IIF) patterns detected during routine antinuclear antibody (ANA) screening. Although various researchers have studied the DFS pattern in recent years, it presents a clinical puzzle since its significance remains elusive. Anti-DFS-70 antibodies occur in healthy individuals with different medical conditions. Unlike the other ANA patterns, they are not associated with systemic autoimmune disease. The present study examined the association of a DFS ANA pattern with the medical conditions of patients who presented to a tertiary care medical center.

Methods
Retrospective chart review of all laboratory values (January 2020 through June 2022) with ANA- and DFS-positive patterns in patients with ANA tests ordered by various clinicians and compared to the patient diagnosis reported. UMass Human IRB approves this study.

Results
Of 8885 ANA samples analyzed, 67.8% (6022) were reported negative. Of total positive ANA (2863) samples, 78.6% (2257) had nuclear patterns, 9.1% (260) had mitotic patterns, and 12.1% (346) had cytoplasmic patterns. Of the positive ANA samples, 14.6% (417) had DFS patterns. For samples with DFS pattern, titers ranged from 1:40 to 1:1280; 62.5% were between1:40, and 1:80. Of samples positive for ANA DFS patterns, 76% (317) were from female patients, and 90% (375) were from patients &gt;21 years of age. Among patients with DFS patterns, the most commonly reported diseases were neurological, followed by respiratory, digestive, circulatory, and musculoskeletal (Table 1).

Conclusion
Clinicians should use a positive ANA with a DFS pattern to indicate pathological phenomena other than autoimmune diseases. Only 3.1% of the patients with DFS patterns have a systemic rheumatic disorder. Clinicians should pay close attention to ANA Patterns, especially DFS staining when using ANA testing.
B-143 Distribution of Uricemia Levels in the Population of a Third-level Hospital. Study of the Prevalence of Hypouricemia
Authors: Corominas A I , Casal G H , Vommaro F M , Riquelme G B , Balconi S M , Damiano A E 
Published: September 2023

Abstract
Hypouricemia is a serum uric acid concentration ≤2.0 mg/dL. The causes may be congenital or acquired. Despite not being recognized as a relevant biomarker, the knowledge of the importance of uric acid in redox homeostasis and the hierarchization of hyperuricemia as an independent risk factor in metabolic diseases justifies a reassessment of the significance of the state of acquired hypouricemia. We aimed to study the prevalence of hypouricemia and its relationship with the referred patients’ ages, genders, and morbidities. A descriptive, quantitative study was carried out, through the survey and analysis of biochemical records and clinical records. We worked with the database of the laboratory’s system between January and October 2019, with the data groups of adult patients (≥16 years) that included serum uric acid dosage. The concomitant biochemical parameters (Urea, creatinine, white blood cell count, glucose) were evaluated, and the diagnoses and treatments were collected in the electronic medical record. To define the prevalence, we worked with the first analysis group of each patient. All statistical analyzes were carried out with the Statistix™ software version 7. The criteria for statistical significance were P &lt; 0.05. We found that of the total 159 971 data groups from January to October 2019, 17 983 were selected. There were 234 patients with hypouricemia analyzed. The prevalence of patients with hypouricemia was 2.22%. In our population, postmenopausal women did not match the distribution of uricemia levels to that of men, nor did the mean values of each distribution. Men had higher levels of uric acid than women in any age situation. The prevalence of hypouricemia agrees with what is described in the literature. Of women with hypouricemia, 39% were in the process of pregnancy. Besides that, most patients had a pathology, less than 10% were present for health control, or did not have data in the clinical record. The prevalence of oncological pathology (46%) as a comorbidity in patients with hypouricemia (oncohaematological tumors more frequent than solid ones) and diabetes (22%) was striking. Hypouricemia values were more frequent in outpatients (70%) than in hospitalized patients, however, the last ones showed lower values (1.46 ± 0.4 mg/dL vs 1.52 ± 0.35 mg/dL). The white blood cell count values (0.1–26 * 103 cells/µL) and glucose (0.3–3.76 g/L) that accompany the hypouricemia situation behold pathological values, and the urea and creatinine values were normal. Because this is a retrospective work, we do not have studies of fractional excretion of uric acid, which could delve into the causal research of hypouricemia. Further studies are needed to elucidate the causes of hypouricemia and to clarify its value in diagnosing and treating diseases.
B-155 Sigma Metric Equation Proposed by James O. Westgard is Wrong
Authors: Brooks Z C 
Published: September 2023

Abstract

Background
2022 AACC Poster #132 of the same title described how the Westgard sigma metric equation of [A] SM = (TEa - Bias)/SD proposed by James O. Westgard is NOT wrong when compared to the industrial practice of calculating sigma as [B] the difference between the mean and nearest upper or lower tolerance limit divided by the SD. Formulae [A] and [C] produce the same results. The often-recommended practice of calculating sigma as [C] Sigma = (%TE -|%Bias|)/CV%, however, produces results that differ from either formulae described above whenever bias is not zero. This formula [C] using percentages is therefore wrong.

Methods
Sigma values were calculated as [A] (TEa - |Bias)|/SD and as [C] (%TE -|%Bias|)/CV% on fourteen routine chemistry analytes and four cardiac markers tested on two Ortho Vitros 5600 analyzers from September to December 2022. CLIA TEa limits were selected as TEa limits for each analyte; bias was calculated as Current Mean - Peer Mean. Sigma tables were used to convert sigma to defects per million results. Eight-six of 320 data sets with sigma metrics from 2.25 to 5.25 were graphed to compare calculated sigma values and #failures/year.

Results
Comparison of sigma metrics calculated from formula [A] and [C] revealed 1. An R² = 1.00 if all 320 values with sigma metrics from 0.71 to 30.8; 2. An R² = 0.92 when the range was truncated to the 86 samples with sigma from 2.25 to 5.25; and 3. An R² value of only 0.75 when the same sigma values are converted to the number of results failing TEa limits per one million patent samples.

Conclusion
Calculations of sigma metrics based on %Bias and %CV are incorrect. This is exacerbated when sigma is converted to the number of patient failures per million results.
B-181 Multi-centre Analytical Performance Evaluation of the EXENTâ Solution for Identification and Quantitation of Monoclonal Immunoglobulins
Authors: Stanton R , McEntee D , Matters D , Ijaz S , Khalid A , Barnidge D , Ouverson L , Sakrikar D , Ashby J , Marrott N , Ghobrial I M , El-Khoury H , Bertamini L , Fleming G , Horowitz E , Neish A S , Solis Z M , McLendon K B , North S 
Published: September 2023

Abstract

Background
Current routine methodologies for monoclonal immunoglobulin measurements may not be sufficiently sensitive to reflect the depth of response seen in patients since the introduction of novel therapies; mass spectrometry may offer a valuable, sensitive alternative approach. Here we describe the preliminary analytical performance characteristics of the EXENT® solution (in development by The Binding Site, part of Thermo Fisher scientific) that combines specific immunoprecipitation steps and mass spectrometry for the identification and quantification of IgG, IgA and IgM intact monoclonal immunoglobulins. Each intact monoclonal immunoglobulin clone can be tracked using its unique m/z value.

Methods
The Lower Limit of Measuring Interval (LLMI) was established for each immunoglobulin type following EP17-A2:2012. Linearity studies were performed according to CLSI EP06-A2:2020 using high and low pools of IgG, IgA, and IgM monoclonal samples and with additional linearity testing below 1 g/L for each specificity to more effectively demonstrate low-end linearity. Within run, between run, between analyzer, between lot and total precision for M protein concentrations and for molecular mass (m/z) of the monoclonal peaks were assessed according to CLSI EP5-A3-2014. Interference was tested following ED3:2018 using 20 potential interferents against 7 samples including high and low IgG, IgA and IgM monoclonal samples.

Results
The initial indication of the EXENT solution performance, in development, are set out below. Results suggest an analytical sensitivity of the assays around 15 mg/L at the LLMI for all specificities. Linearity over a range of 0.014–88.9 g/L for IgG, 0.011–68.4 g/L for IgA, and 0.11–74.2 g/L for IgM. Coefficients of variation (CVs) for M protein concentrations in precision studies were &lt;15% for all specificities and samples. Mass/charge values were within ±1.1 to ±1.5 m/z in total precision studies, and within ±2.7 to ±3.9 m/z in between lot precision studies, respectively, for M proteins with an m/z value ranging from 11 360.6 to 11 698.5 m/z. No significant interference effects were observed when testing the 20 interferents including intralipid (20 g/L), triglyceride (15 g/L), bilirubin (400 mg/L), rheumatoid factor (200 IU/mL) and haemoglobin (10 g/L).

Conclusion
The new EXENT solution demonstrates the potential for a wide measuring interval and the ability to detect M proteins with very low concentration. Also, it could provide stable and reproducible performance for the detection and typing of monoclonal immunoglobulins.
B-208 Correlation Studies of Human Papillomavirus DNA on Roche Cobas vs Human Papillomavirus mRNA on Hologic Panther
Authors: Teng J 
Published: September 2023

Abstract

Background
Molecular testing, specifically HPV DNA has revolutionised the screening and management of cervical cancer. WHO in 2021 has recommended HPV DNA as the primary screening test and also the newer technology HPV mRNA as alternative assay. This evaluation aims to study the result concordance between HPV DNA and mRNA assay.

Methods
The evaluation was conducted on Roche Cobas 4800 (HPV DNA) and Hologic Panther (HPV mRNA) at NHGD IMH lab in 2021. Both instruments underwent precision studies and carryover test to ascertain its performance prior to the concordance study. 64 samples were used in the correlation against the reference method- HPV DNA testing on Roche Cobas 6800.

Results
Both Cobas 4800 and Hologic Panther demonstrated acceptable performance in precision, carryover and LoD (for Hologic only). However closer result agreement is observed in Roche 4800 results to the testing laboratory, Quest which is using Roche 6800.

Conclusion
The disconcordance between cobas 6800 vs cobas 4800 came from the negative samples in cobas 4800 in which their Ct value were found to be near to the detection limit. Similar to Cobas 4800, the discordance observed in Hologic came from the negative samples. However, HPV mRNA screening has been recognized for its higher specificity at baseline in the recent studies which improve clinical outcomes and reduce colposcopy referrals by 14%. Unfortunately, this could not be verified in our evaluation as colposcopy results for those HPV DNA positive sample were not available.
B-215 Use of Whole Exome Sequencing and T Cell Receptor β Repertoires Analysis for Identifying Potential Genetic Variants in Rheumatoid Arthritis
Authors: Cho J , Lee H , Ahn S , Song J , Kim J , Lee J 
Published: September 2023

Abstract

Background
Rheumatoid arthritis (RA) is one of the most prevalent autoimmune diseases, which is reported to affect about approximately 1% of the adult population worldwide. And, it is associated with progressive disease courses, systemic complications, early mortality, and a high socioeconomic burden. The pathogenesis of RA is too complex to be fully understood yet, but abnormal and pathogenic T-cell responses that evade normal immune functions could be considered one of the mechanisms of RA development. Herein, we used whole exome sequencing (WES) and T cell receptor/T cell receptor β repertoires analysis (TCR/TRB) to identify the potential genetic variants in RA.

Methods
Peripheral blood samples from 18 RA patients and 5 healthy controls were analyzed. For RA patients, the baseline naïve samples were collected before disease-modifying anti-rheumatic drugs treatment was started, and both WES and TRB were tested. After 6 months and 12 months of treatment initiation, additional samples were collected and TRB was tested. A total of 18 WES and 27 TRB tests were conducted. WES was done on the MGI-G400 platform (MGI Tech Co., Ltd., Shenzhen, China) and TCR/TRB was done using the LymphoTrack TCR/TRB assay (Invivoscribe Technologies, San Diego, CA, USA) on the MiSeq system (Illumina Inc, San Diego, CA, USA).

Results
In RA patients, deleterious variants that have been reported to be relevant in various diseases were identified in a total of 217 genes. Among 18 RA patients, 5 (27.8%) patients had variants that were suspected to be associated with the pathogenesis of RA, including JAK3, PADI4, TNFSF18, TRAF1, NFKB, etc. Of the 27 TCR/TRB tests, 10 TCR (37.0%) and 14 TRB (51.9%) results had significant changes in RA patients.

Conclusion
The aim of this study is to provide clinical evidence for developing diagnostic and/or prognostic markers for RA. Further analysis whether these variants found in this study are clinically significant and valuable as markers in RA. In addition, it is expected that the results of our study will be used as basic data applied to other rheumatoid diseases such as systemic lupus erythematosus and Bechet’s disease.
B-222 Validation of a qPCR Method for Qualitative Detection of Monkeypox Virus
Authors: Chan A , Amaro K , Bui C , Camarena O , Dang A , Herron D , Lara J , Lim L , Lopez P , Mayoralgo M , Nguyen A , Nguyen J , Ortega B , Pham A , Tran K , Bach D 
Published: September 2023

Abstract

Background
The 2022 Monkeypox virus (MPXV) outbreak became a global concern, prompting a multidisciplinary response to increase surveillance and isolate suspected and confirmed cases to reduce disease spread. Quantitative PCR (qPCR) is capable of detecting small amounts of DNA or RNA, and can be used to quickly and accurately confirm MPXV cases, even at early stages of infection, which is crucial for outbreak control and management.

Methods
qPCR was performed on a 384-well plate with a total reaction volume of 20 μl consisting of 9 μL Acrometrix Monkeypox DNA Positive Control at serial dilutions, 10 μL BactoPure Master Mix, and 1 μL TaqMan Monkeypox Virus Microbe Detection Assay. Positive control dilutions 103, 102, 101, and 100 were diluted with TE Buffer. qPCR was performed on two different Quantstudio 12K Flex instruments.

Results
The qPCR method validation was performed by calculating precision, accuracy, and linearity on serial dilutions of Acrometrix Monkeypox Virus DNA Positive Control. Precision testing was performed on 2 levels of control: low (101) and high (103) over 5 days with 4 repeats per day. Accuracy and linearity testing were performed on 4 levels with 3 repeats. Coefficient of variation (CV) was less than 5%, specifically 0.9% for the low control and 4.3% for the high control, indicating an acceptable level of precision. Percent recovery fell close to 100%, specifically within 101%-103% for all controls, indicating the results fell close to the expected value. Linearity R2 was close to 1, specifically 0.984, demonstrating the results are proportional to the concentration analyzed.

Conclusion
Validation of MPXV detection through qPCR was successful. Development and validation of qPCR assays for the detection of positive MPXV cases is essential for managing future outbreaks and combatting future pathogenic strains.