Clinical Chemistry Analyzer Research Articles

Comparative Study of Subclinical Hypothyroidism in Patients with and Without Metabolic Syndrome

Background: A group of metabolic abnormalities known as metabolic syndrome (MetS) greatly raises the risk of type 2 diabetes mellitus and cardiovascular illnesses. People with MetS frequently have subclinical hypothyroidism (SCH), which is defined by high levels of thyroid-stimulating hormone (TSH) with normal free thyroxine (T4). The research examined the relationship between thyroid function and metabolic parameters, as well as to assess the incidence of subclinical hypothyroidism in individuals with and without MetS. Methods: There were 960 participants in all, 480 of whom had a diagnosis of MetS and 480 of whom were healthy controls. Blood pressure and waist circumference were measured clinically. After an overnight fast, blood samples were taken utilising fully automated clinical chemistry and hormone analyzers to assay lipid profile, fasting blood glucose (FBG), and thyroid function tests (T3, T4, and TSH). The Pearson correlation coefficient and Student's t-test were used to analyse the data. Results: Statistically, there was a significant variation in the prevalence of subclinical hypothyroidism between the MetS group (16.3%) and the control group (5%; p < 0.001). In addition to having decreased HDL cholesterol, MetS patients also had higher FBG, blood pressure, triglycerides, and waist circumference. TSH levels and the elements of MetS, such as waist circumference (r = 0.41, p < 0.001), systolic blood pressure (r = 0.38, p < 0.001), FBG(r = 0.44, p < 0.001), triglycerides (r = 0.39, p < 0.001), and HDL cholesterol (r = -0.32, p < 0.001), were found to be significantly correlated with each other. Conclusion: The results of the study indicate that people with metabolic syndrome have a higher incidence of subclinical hypothyroidism, which may indicate a relationship between thyroid function and metabolic health. Regular thyroid function testing could help improve clinical outcomes by enabling early detection and management of SCH in MetS patients. Recommendations: When treating patients with metabolic syndrome, healthcare professionals should think about doing routine thyroid function tests. To manage thyroid dysfunction in patients with metabolic syndrome (MetS), further research is required to investigate the underlying mechanisms and create tailored therapies. Keywords: Metabolic Syndrome, Subclinical Hypothyroidism, Thyroid Function, Cardiovascular Risk, Metabolic Health

Study of an outbreak of jaundice in Rajshahi, Bangladesh: a sociodemographic and clinical manifestation

BackgroundJaundice symptom outbreaks occur very frequently in Rajshahi, Bangladesh, from January to March 2024. The present study was designed to verify the pattern of jaundice outbreaks and identify the affected population and treatment of jaundice patients.MethodsA total of 1151 students living in Rajshahi were tested as samples. Jaundice syndrome was determined by physiological and laboratory tests. Blood and urine samples were tested via an automated clinical chemistry analyzer to confirm the pathophysiology. Moreover, sociodemographic status, food habits and living areas were considered for the patients.ResultsAmong the total number of tested patients, 27.28% and 72.72% were jaundice positive and negative, respectively. In the sociodemographic study, the highest percentage (94%) of jaundice-positive patients were men, which was found in patients aged 20–30 years. In the case of the clinical test, the jaundice-positive patients were hepatitis A virus (HAV)-positive, and the highest average alanine transaminase (ALT) and total bilirubin levels were 1312.67 ± 1.32 U/L and 2.08 ± 0.70 mg/dL, respectively.ConclusionThe findings of the present study revealed that the jaundice outbreak was caused by HAV. This study may be helpful for effectively measuring jaundice outbreaks to ensure that safe food and drinking water are urgently needed.

Towards routine high-throughput analysis of fecal bile acids: validation of an enzymatic cycling method for the quantification of total bile acids in human stool samples on fully automated clinical chemistry analyzers.

Bile acid diarrhea is a common but underdiagnosed condition. Because the gold standard test (75SeHCAT) is time-consuming and not widely available, fecal bile acid excretion is typically assessed by chromatography and mass spectrometry. Although enzymatic cycling assays are well established for the rapid and cost-effective analysis of total bile acids (TBA) in serum or plasma, their full potential has yet not been extended to stool samples in clinical routine. The performance of the 'Total bile acids 21 FS' reagent (DiaSys) was evaluated in fecal matrix according to CLSI guidelines and EU-IVD Regulations (2017/745), and compared to an established microplate-based kit (IDK®) by measuring patient stool samples (n=122). Method agreement was assessed by Passing-Bablok and Bland-Altman analysis. The quantification of eight individual BAs was assessed using HPLC-MS/MS as reference method. The DiaSys assay showed linearity between 3.5 and 130 μmol/L, good repeatability, total precision, and reproducibility with CVs of 1.7 %, 3.5 %, and 3.0 %. Limit of blank (LoB), detection (LoD), and quantitation (LoQ) were≤0.17,≤0.3, and 3.5 μmol/L, respectively. No significant interference from endogenous substances was observed. The methods showed good correlation up to 140 μmol/L (r=0.988), despite differences in the quantification of individual BAs, with mean deviations of 7 % (DiaSys) and 31 % (IDK®), respectively. The advantages of enzymatic TBA analysis on fully automated clinical chemistry platforms can be exploited for the routine analysis of stool samples. However, cycling assays may benefit from reference standards that take into account the composition of the fecal BA pool.

Study of Deritis Ratio in Acute Myocardial Infarction Patients in a Tertiary Care Hospital

Introduction: The ratio of Aspartate Aminotransferase (AST) to the Alanine Aminotransferase (ALT) is known as the Deritis ratio. Many studies have highlighted, the significance of AST and ALT measurement as a prognostic significance in cardiovascular diseases including acute myocardial infarction. The current study aimed to discuss the correlation of Creatine Kinase MB (CKMB) and Cardiac Troponin I with the Deritis ratio in the retrospectively enrolled Nepalese population. Materials and Methods: Two hundred and sixty-four (264) cases of acute myocardial infarction who had undergone AST and ALT measurement were enrolled for the study by non-probability purposive sampling technique. The study variables are AST, ALT, CKMB, Troponin I, age, etc. The above-mentioned variables were measured by using a fully automatic clinical chemistry analyzer Beckmann Coulter AU480 after proper quality control check and calibration. Results: Out of 264 patients, 162 were male and 102 were female. There was no significant difference in age (P value: 0.54) and Deritis ratio (P value: 0.73) between the male and female genders when the mean and standard deviation were compared. The non-parametric correlation (Spearman’s rho) analysis revealed the positive correlation of Deritis ratio with the cardiac troponin I (0.18, 0.00) which is statistically significant. Secondly, the Deritis ratio was positively correlated with the CKMB (0.40, 0.00) which was statistically significant. Conclusion: Our results have shown the raised Deritis ratio in acute myocardial infarction patients, which represents the release of AST during the myocardial infarction or the stress to the hepatocytes due to the hemodynamic changes occurring during myocardial infarction.

Evaluation of the Accuracy of FreeStyle Libre 2 for Glucose Monitoring in White New Zealand Rabbits.

Studies are currently being conducted on rabbits requiring serial glucose monitoring. The FreeStyle Libre 2 (FSL2), a serial glucose monitoring device, has been validated in humans, dogs and cats, but not in rabbits. This study aimed to evaluate the accuracy of the FSL2 in rabbits. Six healthy rabbits were used in this study. Interstitial glucose (IG) was measured using the FSL2, and blood glucose (BG) was measured using a portable blood glucose meter (PBGM); their results were compared with those from a clinical chemistry analyser. For the first 3h, IG and BG were measured at 1-h intervals. Subsequently, they were measured every 8h over a 48-h period. Regular insulin 0.2 U/kg was then administered to the rabbits, and IG and BG were measured every 15min over a 90-min period. Before insulin treatment, no measurements fell within the hypoglycaemic range (BG < 100mg/dL). In the euglycaemic range (BG ≥ 100mg/dL), the PBGM and FSL2 showed 85.7% and 23.8% accuracies, respectively. After insulin treatment, the PBGM showed 95.5% and 81.3% accuracies in the hypoglycaemic and euglycaemic ranges, respectively. The FSL2 showed 68.1% and 37.5% accuracies in the hypoglycaemic and euglycaemic ranges, respectively. Parkes consensus error grid analysis showed that the PBGM and FSL2 had 100% agreement for Zones A (no effect on clinical action) and B (altered clinical action unlikely to affect outcome) in rabbits with and without insulin treatment. There was limited agreement between the FSL2 and reference standard BG measurements. However, the FSL2 allows clinically acceptable identification of hypoglycaemic states in rabbits.

Comparative Study of Human Serum Paraoxonase-1 (PON-1) Activity in Type 2 Diabetes Mellitus: Insights into Oxidative Stress and Antioxidant Defence

Diabetes mellitus is a disease characterized by chronic high blood sugar levels resulting from defects in insulin secretion, insulin action, or both. Prolonged high blood glucose can increase production of reactive oxygen species, leading to elevated oxidative stress. The enzyme paraoxonase-1 (PON1), made in the liver and transported on high-density lipoprotein (HDL), acts as an antioxidant and can counter oxidative stress. This study aimed to compare PON1 activity between healthy controls, obese type 2 diabetics, and non-obese type 2 diabetics. There were 30 healthy controls, 30 obese type 2 diabetics (group I), and 30 non-obese type. Additionally, fasting lipid profiles, fasting plasma glucose, uric acid, albumin, and total bilirubin levels were determined using a clinical chemistry analyzer. The goal was to evaluate differences in PON1 activity as well as metabolic parameters between the three study groups. The results In comparison to the healthy control group, the diabetes group had Significantly higher blood sugar, serum cholesterol, serum triglycerides, serum LDL, and serum uric acid levels, Compared to the healthy control group, PON was Significantly lower in group II patients, Conclusion that Obese type 2 DM patients had significantly lower PON1 and HDL-C levels, which may indicate That the overweight has lessened biochemical functions for these substances. As dyslipidemia, insulin resistance, and high blood pressure are thought to be crucial factors in the cause of metabolic conditions in obese people, the reduced paraoxonase level may increase their likelihood of developing these conditions.

Canonical Correlation of Milk Composition Parameters and Blood Biomarkers in High-Producing Dairy Cows During Different Lactation Stages.

This study explores milk composition and blood markers in cows across lactation stages. Holstein cows were divided into four groups: beginning of lactation (BL; n = 21), peak of lactation (PL; n = 21), middle of lactation (ML; n = 21), and end of lactation (EL; n = 20). Blood (1 × 15 mL) and milk samples (1 × 100 mL) were collected for biomarker analysis. Blood chemistry profiles were determined using a clinical chemistry analyser, and milk lactose, fat, and protein levels (%) were determined using an infrared absorbance analyser. Minerals (Ca, P, and Mg) in milk were determined by atomic absorption spectrometry after mineralizing the samples. Glucose was higher in the EL group than in the BL group (p < 0.01), whereas D-beta-hydroxybutyrate (D-BHB) was higher in the BL group than in the PL and ML groups (p < 0.001). Cholesterol was higher in the PL, ML, and EL groups than in the BL group (p < 0.001). Gamma-glutamyl transferase was increased in the PL group compared to the BL group. Phosphorus levels were lower in the PL than in the BL group, whereas protein levels were higher in the EL than in the PL group. Spearman and partial correlation analysis showed several significant associations between the observed variables. Using canonical correlation analysis were identified three significant correlations (rc1 = 0.853; rc2 = 0.823; rc3 = 0.739). The main canonical correlation identified blood TG and milk urea as the strongest variables. According to the canonical loading, the biomarkers TG, Mg, urea, cholesterol, and alkaline phosphatase (U1) are the primary variables associated with milk parameters (V1), specifically with milk urea, milk Mg and P, protein, and lactose.

Cover, Content, and Editorial Note from Borneo J Med Lab Tech Vol. 7 No. 1 Oktober 2024

Assalamu’alaikum Wr. Wb. Alhamdulillahirabbil ‘alamin. After a long wait of almost 12 years from the first stand, the scientific journal of the Department of Analyst Health (Medical Laboratory Technology) Universitas Muhammadiyah Palangkaraya can be published. BJMLT is published every 6 months (2 issues/year) every October and April. This journal aims to publish high-quality articles dedicated to all aspects of the latest outstanding developments in the field but not limited to Blood transfusion, Clinical Chemistry, Hematology, Histopathology, Immunology, Microbiology, Parasitology, Toxicology, Food and Beverage Analysis, Molecular Biology, Medical Biology, and related fields that are of interest to medical laboratory technologies, such as epidemiology, health promotion, professional ethics, and laboratory management, It was first published online in October 2018 using Open Journal System (OJS) 3.1 platform with e-ISSN 2622-6111 by URL http://journal.umpr.ac.id/index.php/bjmlt. This edition contains 14 articles on Hematology, Microbiology, Clinical Chemistry, and Epidemiology. Editorial boards are fully aware that there is still room for improvement in this edition; hence with all humility, they are willing to accept constructive suggestions and feedback for improvements to the publication for the next editions. The publication of BJMLT certainly participates in disseminating the results of research and review of science and technology development conducted by lecturers and researchers from UM Palangkaraya and other universities. The editorial board would like to thank the University, all editors and reviewers, and contributors of the scientific articles who have provided the repertoire in this issue. We hope that all parties, especially the contributors of the articles, could re-participate the publication's success in the next edition in October 2024.

7872 Adiponectin Enhances Hepatic Insulin Sensitivity in a Rat Model of Polycystic Ovary Syndrome: Role of PI3K/Akt/mTORC2 Pathway

Abstract Disclosure: A. Abdelhameed: None. S. Rezq: None. R.M. Vinson: None. J. Flaherty: None. B. Kim: None. D.G. Romero: None. L.L. Yanes Cardozo: None. Background: Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in reproductive-age women and is associated with a high prevalence of obesity and insulin resistance (IR). Phosphoinositide-3-kinase (PI3K)/Akt/ the mammalian target of rapamycin complex 2 (mTORC2) pathway is one major insulin signaling pathway and its impairment is involved in multiple metabolic disorders by inducing IR. Phosphorylation of protein kinase B (Akt) at (Ser)-473 by mTORC2 leads to its full activation and subsequently enhances insulin sensitivity. Adiponectin is a peptide secreted by adipocytes that increases insulin sensitivity. PEG-BHD1028 is a novel potent peptide adiponectin receptor agonist that enhances insulin sensitivity in different models of diabetes. However, its potential to attenuate androgen-mediated hepatic IR by modulating PI3K/Akt/mTORC2 pathway remains unknown in PCOS. Hypothesis: We tested the hypothesis that PEG-BHD1028 attenuates hepatic IR via modulating PI3K/Akt/mTORC2 pathway in a rat model of PCOS. Methods: To induce PCOS, four-week-old female Sprague Dawley rats were implanted with dihydrotestosterone (DHT) silastic tubes (7.5mg) or placebo (PBO) for 90 days (n=4-8/group). During the final three weeks of DHT treatment, rats received PEG-BHD1028 (25μg/kg/day) subcutaneously. Body weight (BW) by gravimetry, fat and lean mass were measured by Echo-MRI. Fasting plasma insulin was measured by ELISA while fasting plasma glucose was measured with a clinical chemistry analyzer. HOMA-IR, an index of systemic IR, was calculated. Protein levels of insulin signaling markers including IRS1, p-IRS1 (Ser1101), PI3 kinase p110, Akt, p-Akt (Ser473), mTOR, p-mTOR (Ser2481) and mTORC2 regulatory protein Rictor were quantified in the liver by Western-blot. Results: DHT significantly (p&amp;lt;0.05) increased BW, fat mass and lean mass by 1.4-, 2.2-, 1.4- folds, respectively compared to PBO group. DHT significantly (p&amp;lt;0.05) increased HOMA-IR by 3.8- fold, compared to PBO group. DHT downregulated hepatic p-Akt, Akt and mTOR protein expression by 20% to 40% while upregulating IRS-1 by 1.6- fold with no impact on PI3 kinase, p-mTOR, Rictor and p-IRS1 compared to the PBO group. PEG-BHD1028 significantly decreased BW, fat mass and lean mass while attenuating IR (34.7% reduction in HOMA-IR) in PCOS rats. Interestingly, PEG-BHD1028 significantly (p&amp;lt;0.05) increased PI3K, active Akt (pAKT), mTOR and Rictor levels by 1.6-, 2.2-, 1.5-, 1.8- folds, respectively with no effect on the other insulin signaling markers compared to vehicle-treated group. Conclusion: Our findings suggest that PEG-BHD1028 is a novel and effective therapeutic agent to attenuate hepatic IR in PCOS via activating PI3K/Akt/mTORC2 pathway. Significance: Targeting adiponectin signaling could be a novel and effective therapeutic approach to mitigate metabolic dysfunction in females with excess androgens. Presentation: 6/2/2024

B-039 Evaluating Inter-Analytic Module Immunoassay Carryover

Abstract Background Integrated analytical systems contain both clinical chemistry analyzers and immunoassay analyzers. Immunoassays can be assigned high priority based on potential sample carryover from sample probes used in clinical chemistry analyzers. The priority designation, assigned by the vendor, determines if the immunoassay test (disposable pipette) is sampled prior to the clinical chemistry test (non-disposable pipette). However, in a modern-day core chemistry laboratory, add-on testing is prevalent. High priority assignment by analytical systems is only effective when a specimen’s tests are analyzed concurrently and doesn’t account for an add-on of any high priority immunoassays. Evaluation of carryover for high priority immunoassays on clinical chemistry analyzers provides insight on if there is clinical impact of allowing add-on testing of these analytes. Methods Intact human chorionic gonadotropin + β-subunit (HCG-BETA, Roche Diagnostics, cobas e602), Procalcitonin (PCT, Roche Diagnostics, cobas e602), and Elecsys Total Prostate-specific Antigen (TPSA, Roche Diagnostics, cobas e602) were the high priority immunoassays evaluated for carryover. High and low target pools were separated into aliquots and baselines for each pool established. High sampling volume assays to potentially generate carryover on the clinical chemistry analyzers were determined to be Sodium (Na, Roche Diagnostics, cobas ISE), Unsaturated Iron-Binding Capacity (UIBC, Roche Diagnostics, cobas c702), and Ammonia (NH3L2, Roche Diagnostics, cobas c502). The ordering sequence of high pool to low pool was then assayed in triplicate on each clinical chemistry analyzer. After potential carryover had been generated, the low pool was again assayed for the immunoassays to evaluate if carryover had occurred and if the carryover exceeded the analyte’s allowable error. Statistical analysis was then performed utilizing Microsoft Excel. Results Conclusions Add-on testing should be restricted for β-HCG and TPSA as a previous clinical chemistry analysis could lead to an errant add-on result due to carryover. PCT is unaffected by previous clinical chemistry analysis.

B-008 An Evaluation of the Analytical Performance of the New Beckman Coulter DxC 500i Clinical Analyzer

Abstract Background The DxC 500i Clinical Analyzer* is Beckman Coulter’s latest integrated Clinical Chemistry and Immunoassay analyzer. It provides reliability, scalability, and workflow efficiency via a flexible independent maintenance mode, increased uptime, and a dynamic sample handler to optimize sample management. Standard Beckman Coulter Chemistry and Immunoassay consumables and reagents ensure network standardization and commutability. The purpose of this study was to evaluate the analytical performance of the new DxC 500i analyzer and to compare the performance against the Beckman Coulter stand-alone Clinical Chemistry and Immunoassay analyzers on the market. Methods To assess performance over a range of assay methodologies, several DxC 500i Beckman Coulter Immunoassays and Chemistry assays were evaluated. Precision was assessed using human samples following CLSI EP05-A3 guidelines. Linear range was assessed following CLSI EP06-A guidelines. Method Comparison and Bias Estimation studies compared the DxC 500i Clinical Analyzer against the Access 2 Immunoassay System, the AU480 and the DxC 700 AU Clinical Chemistry analyzers, following CLSI EP09c-ED3 guidelines. To investigate carryover potential as samples are processed across the integrated system, a carryover study was designed. This focused on evaluating potential sample-to-sample carryover caused by the AU sample probe and its effects on immunoassay samples during reflex scenarios. Results DxC 500i Chemistry results: Glucose Serum (OSR6x21) Assessments of repeatability and within laboratory precision at multiple critical analyte concentrations showed total imprecision of &amp;lt;3% and within-run imprecision of &amp;lt;3%. The Glucose serum assay demonstrated acceptable linearity throughout the 10-800 mg/dL analytical measuring range. Method comparison studies compared the DxC 500i analyzer against the AU480 and DxC 700 AU systems. A Weighted Deming regression of serum patient samples (n&amp;gt;100 measured across the analytical range) yielded a 0.986 slope, 0.227 mg/dL intercept, and R=0.9999 correlation coefficient against the DxC 700 AU and a 0.995 slope, 0.356 mg/dL intercept, and R=0.9999 correlation coefficient against the AU480. DxC 500i Immunoassay results: Cortisol (33600) Beckman Coutler’s Cortisol assay exhibited total imprecision of &amp;lt;12%CV at &amp;lt;5 μg/dL or &amp;lt;10CV% at ≥5 μg/dL for serum. The Cortisol assay demonstrated acceptable linearity throughout the 0.4-60 μg/dL analytical measuring range. A method comparison study compared the DxC 500i against the Access 2 system. A Weighted Deming regression of serum patient samples (n&amp;gt;100 measured across the analytical range) yielded a 0.9785 slope, 0.2764 μg/dL intercept, and R=0.9932 correlation coefficient. DxC 500i Carryover Assessment The Total βhCG (5th IS) assay was used to evaluate the potential of chemistry analyzer to immunoassay analyzer sample carryover. The study used High hCG serum samples (&amp;gt;1,000,000 mIU/mL) and Low hCG serum samples (4-6 mIU/mL). The DxC 500i exhibited a cumulative carryover of ≤2 parts per million (PPM) for serum. Conclusions These studies demonstrate excellent analytical performance for the new Beckman Coulter DxC 500i Clinical Analyzer* and confirm comparable performance to current Beckman Coulter standalone Clinical Chemistry and Immunoassay analyzers on the market. *Pending clearance by the United States Food and Drug Administration; not yet available for in vitro diagnostic use in the U.S. Product availability and regulatory status depends on country registration per applicable regulations.

B-295 Sensitive and Rapid Homogeneous Immunoassay for the Detection of Zolpidem and its Major Metabolite in Urine

Abstract Background Zolpidem, a schedule IV controlled substance under the Controlled Substances Act, is commonly known as Ambien and Ambien CR, a prescription only treatment for insomnia. Zolpidem is one of the most common prescribed sleep aids in the U.S. due to its rapid sedative effects. The recommended duration of treatment is between two days and four weeks, as long-term use of zolpidem increases the risk of habit formation. Addiction to zolpidem can cause severe side effects, including memory loss, delusion, and in extreme cases, fatal overdose. From 2005-2010, Zolpidem-related emergency room visits doubled, underscoring the need for accurate testing as zolpidem testing plays an important role in cases of misuse, diversion, and forensics. The sole commercially available homogeneous immunoassay for the detection of zolpidem in urine has a relatively high cutoff at 20 ng/mL and poor cross-reactivity (0.02%) to major metabolite zolpidem 4-carboxylic acid. ARK Diagnostics has developed the ARK™ Zolpidem Assay to detect zolpidem at a cutoff concentration of 10 ng/mL with high cross-reactivity to its metabolite, Zolpidem phenyl 4-carboxylic acid and no cross-reactivity to zaleplon and zopiclone at concentrations below 100,000 ng/mL. Methods The ARK™ Zolpidem Assay is a liquid stable homogeneous enzyme immunoassay, consisting of two reagents, with a cutoff concentration of 10 ng/mL and semi-quantitative range up to 40 ng/mL. Preliminary performance characterization for this assay was evaluated on the Beckman Coulter AU680 Automated Clinical Chemistry Analyzer. Precision, analytical recovery, specificity, Histogram Overlap Analysis of ±50% controls and the cutoff, and method comparison with LC-MS/MS were evaluated. Results In semi-quantitative mode, total precision ranged from 2.6 to 3.1% CV. Spiked recovery of zolpidem ranged from 88.6% (2.5 ng/mL) to 96.8% (10 ng/mL). The major metabolite, zolpidem phenyl-4-carboxylic acid, showed an approximate equivalence to the 10 ng/mL zolpidem cutoff at 30 ng/mL (33.3% cross-reactivity). Histogram overlap analysis showed no overlap between cutoff and control levels. Method correlation with LC-MS/MS using authentic urine samples showed an excellent agreement with a specificity of 100% and sensitivity of 100% (15 positives and 100 negatives). Conclusions The ARK™ Zolpidem Assay measures zolpidem and its major metabolite zolpidem phenyl-4-carboxylic acid in human urine with acceptable performance. The assay is sensitive, rapid, and applicable to a wide range of clinical chemistry analyzers.

B-324 Pilot Study Result of EQA for Fecal Immunochemical Test for Hemoglobin Using a Novel Ready-to-Use Sample Material

Abstract Background The primary screening method for colorectal cancer is fecal immunochemical test (FIT) detecting human hemoglobin (Hb) in feces. There are several tests on the market both for automated analyzers and point-of-care testing (POCT). To ensure high quality results, the tests should be monitored with appropriate quality assessment procedures including participation in external quality assessment (EQA) programs. By participating in an EQA program using a stool-like material, clinical laboratories and POCT sites can fulfil the ISO 15189 requirements of monitoring the complete laboratory process from pre- to postanalytical phases in addition to the quantitative analytical phase of FIT. Here we present the first results of an international EQA pilot study for quantitative FIT using a novel ready-to-use stool-like material, performed by EQA provider Labquality. Methods 40 participants from 8 countries using 9 different FIT reagents were recruited for the pilot study performed in 2023. A set of artificial stool samples containing human hemoglobin uniformly was shipped to the participants at ambient temperature. The sample material is developed by HECTEF (Health Care Technology Foundation, Japan). The participants were instructed to collect the sample with their test specific sample collection kit and perform analysis according to the manufacturer’s instructions as a regular patient sample. Quantitative results (µg Hb/g feces) and qualitative interpretations (positive/negative) were reported in Labquality’s electronic platform LabScala. The quantitative target value was a method group consensus mean. A questionnaire regarding the usability of the sample material was sent to all participants. Results Sample S001 contained no Hb and all participants (40/40) reported a result corresponding to the lower detection limit of their test in use or a low numeric result most likely explained by unspecific background. All reported a negative interpretation. Sample S002 was a high positive sample for Hb. The measured value exceeded the upper detection limit of 4/9 tests (Exdia iFOB, Precision Biosensor; Fecal Occult Blood (iFOB) Neo, Boditech; FOB Test, VedaLab; NADAL FOB Quant test, Nal von Minden). The mean values and CV% were 116 µg/g and 8% for iFOBT, Aidian; 192 ug/g and 4% for Standard iFOB FIA, SD Biosensor; 144 µg/g and 45% for FOB assay kit FOB Gold, Sentinel Diagnostics (different clinical chemistry analyzers used); 91 µg/g and 26% for OC-Sensor FIT, Eiken Chemical. All participants responded a positive qualitative clinical interpretation. The majority of the responders on the feedback questionnaire (26/35) found the samples to be easy to use including comments of the sample being user-friendly and ready-to-use, resembling a patient sample and easy to apply with the test specific sample collection kit. Conclusions The fecal-like sample used in this Labquality’s EQA pilot study evaluates the complete analytical process including the preanalytical, analytical and postanalytical phases. An EQA scheme using this sample material is more compliant to ISO 15189 requirements compared to schemes where liquid or lyophilized samples are used. The new sample material used on this pilot study was positively received by most of the participants and it was compatible with all tests included in the study.

B-032 Evaluation of Assay Performance for Atellica CI 1900 analyzer in Routine Chemistry, Division of Laboratory Medicine, UMMC

Abstract Background The Atellica® CI 1900 (Siemens Healthiness) is a stand-alone, integrated clinical chemistry and immunoassay analyzer. Method verification has been done on this analyzer for 14 parameters based on the guideline of the Clinical and Laboratory Standard Institute (CLSI). Methods The method evaluation protocol consisted of determination of imprecision based CLSI EP 15-A3; CLSI EP06-A2 protocol was used for verification of linearity and method comparison was carry out in between Atellica® CI 1900 with our current Atellica® Solution based on CLSI EP09-A2 protocol. Method comparison data was analyzed by using statistical software, MedCalc. The data were used to calculate bias for Sigma metrics determination. Results Total of 14 analytes including sodium, potassium, chloride, creatinine, urea, alkaline phosphatase, total bilirubin, triglyceride, uric acid, glucose, thyroid stimulating hormone, free-thyroxine, human chorionic gonadotropin and carcinoembryonic antigen were tested. The results showed the repeatability and total imprecision were within acceptable limit as per manufacturer claims for all analytes. It was demonstrated that the analytical system operates within the claimed linearity ranges as stated in the information for use (IFU). Passing - Bablok analysis show the bias for all analytes were within our laboratory pre-set quality specification. The intercept and slope with 95% CI for Passing-Bablok analysis as shown in table 1. Sigma metrics were determined for each assay using the coefficient variation determined from precision study and bias obtained from the method comparison study. It showed that the Sigma for 14 analytes ranging from 3 to more than 6 with more than 93% assay has sigma scores more than 4. Conclusions Atellica® CI 1900 shows acceptable precision and bias for all analytes that evaluated. The analyser is fully comparable with Atellica® Solution. We conclude that Atellica® CI 1900 demonstrates good performance capabilities, making this analyzer suitable for low-to-medium volume laboratory.

B-023 Workflow evaluation of the new Siemens Atellica CI Analyzer

Abstract Background The Atellica® CI Analyzer, Model 1900 is an integrated clinical chemistry and immunoassay analyzer that has the same user interface, general workflow, and planned test menu to that of the larger Atellica® Solution. Atellica CI Analyzer also shares the same reagents and consumables as the Atellica Solution. The purpose of this study was to evaluate workflow performance of the Atellica CI Analyzer in a routine-like clinical laboratory setting. Methods The study was conducted at four international laboratory facilities over a four-month period. Basic throughput of the Atellica CI Analyzer was evaluated at each site by testing quality control (QC) and remnant anonymized patient samples using an 8:2 mixture of chemistry (ion selective electrolytes [IMT], clinical chemistry analytes) and immunochemistry assays. Additionally, the impact of interrupting routine testing for STAT high-sensitivity cardiac troponin samples was also evaluated. Daily, weekly, and monthly maintenance activities were observed, and data collected through direct observations, time and motion studies, and system run logs. Results The Atellica CI Analyzer aspirated patient samples in as little as two minutes (mins) with the time to first result for the mixed assay worklist ranging from 1 to 12 mins from the time of first barcode read. Across the four sites, the time to last result for the mixed assay worklist (average n=495 test results per site) ranged from 52 to 65 mins. The average turn around time (TAT) for high-sensitivity troponin was 9 mins from barcode read. Daily maintenance and QC activities for the Atellica CI Analyzer are comprised of both automated and manual activities. The average daily maintenance hands-on time was 3.6 min, ranging from 1.6 to 6.4 mins across the four sites. Daily QC preparation required &amp;lt;5 mins of hands-on time. The total average hands-on time to complete weekly maintenance for the Atellica CI Analyzer was 7.5 min. Conclusions The Atellica CI Analyzer demonstrated consistent system performance across all four study sites. Implementation of the Atellica CI Analyzer has the potential to increase laboratory efficiency given the minimal hands-on time required for daily and weekly maintenance activities and short TATs.

B-258 Rapid clozapine blood levels on the Beckman Coulter DxC 500 AU

Abstract Background Clozapine is the most effective drug for treatment-resistant schizophrenia. Multiple guidelines strongly recommend clozapine therapeutic drug monitoring (TDM). Measuring clozapine blood levels provides essential information when there are questions of medication adherence, efficacy, toxicity, medication interactions, or concern over other factors that may influence clozapine levels. Turnaround time is essential for effective clinical decisions. However, with liquid-chromatography/mass spectrometry, clozapine levels can take days to report results. To address the medical need for a faster turnaround time, a rapid clozapine immunoassay was developed for automated clinical chemistry analyzers. The immunoassay measuring range is based on established APA Practice Guidelines and ASCP/AGNP Consensus Statement. We developed an analyzer application for the DxC 500 AU (time to first result &amp;lt; 9 minutes) to expand the availability of rapid clozapine blood level testing. Methods The performance of the FDA-cleared clozapine assay was evaluated on the DxC 500 AU according to CLSI guidelines. Precision testing was performed with three clozapine spiked pools over five days with five replicates per sample. Precision sample recovery was assessed to their theoretical spike concentration. Linearity was verified with five spiked serum samples covering the measuring range. Method comparison was performed with 40 remnant patient samples vs. the Beckman Coulter AU480. Results Repeatability CV was ≤5.5% and within-laboratory precision CV was ≤6.1%. Individual deviation was ±20% and mean deviation was ±2% for all samples. Deviation from linearity was &amp;lt;3%. Deming regression statistics for the method comparison were a slope of 0.987, an intercept of 5.7, and an R of 0.9948. The methods are statistically equivalent: the 99% confidence limits contained 1 and 0 for the slope and intercept respectively. Conclusions The precision, recovery, linearity, and method comparison results for clozapine on the DxC 500 AU demonstrate that the analyzer can be used to generate robust results for clozapine.

A-096 A New Direct Bilirubin (DBIL) Assay with Enhanced Performance in Anti-hemolysis Interference on Mindray BS-2800M Analyzer

Abstract Background Serum bilirubin could be used to assess liver functions. Due to clinical importance, it is highly desired to have an accurate bilirubin test. However, there are circumstances where error could occur in bilirubin test results. In particular, the hemolysis interference is one of the major problems that could mislead the direct bilirubin results because of their relatively high occurring frequencies in the clinical laboratories. To resolve this issue and improve the accuracy for the direct bilirubin, we developed a new reagent formulation for DBIL assay that minimizes hemolysis interference. In addition, this reagent also shown good performance characteristics on Mindray BS-2800M Clinical Chemistry analyzer. Methods The new DBIL assay is a vanadate-oxidation method that bilirubin is gradually transformed to biliverdin because of oxidizing agent vanadate, resulting in decreasing of absorbance at 450 nm. Two-level calibration is used for this assay. DBIL concentrations are obtained from the calibration line with signals from the patient samples in the assay. Following CLSI protocols, basic assay performances were evaluated: interference, precision, linearity, method comparison and low end detection (LOB, LOD, LOQ). Results Hemolysis interference was tested in parallel with DBIL reagents from different IVD companies, Roche, Wako and Chinese domestic main-player companies, A, B, and C. The results showed that the reagents of A and B companies were significantly suffered from hemolysis interference with maximum bias exceeded -30%. Although there are some effects on the anti-hemolysis with DBIL reagents of Wako and C company, the DBIL reagent of Mindray was the best with less than -8% of impact when hemolysate hemoglobin was up to 1000 mg/dL (H index). The Diazo DBIL assay from Roche shown issue with hemolysis interference. The interference to our new DBIL assay from lipid (up to 1000 mg/dL) and ascorbic acid (up to 30 mg/dL) were also with less than ±10% impact. The precision study was run according to EP05A3 with two commercial controls (∼ 10 and 30 µmol/L) and three serum sample pools (∼ 6.8,17.1 and 342 µmol/L). The CVs of repeatability were ranged from 0.54% to 0.58% for controls and 0.24% to 0.88% for serum pools, respectively. The within-lab precision was below 2.5%. The analytical linearity was determined as from 0.66 µmol/L to 439.66 µmol/L. For low-end detection, the LOB, LOD and LOQ were 0.5,1.0 and 1.7 µmol/L, respectively. In addition, the Mindray (y) correlated well with the DBIL assay of Wako (x) reagent on Mindray BS-2800M system (y): y =0.9847 x+0.1301 (r =0.9997, n =400, sample range: 1.08 - 430.00 μmol/L). Conclusions The new Mindray direct bilirubin assay shown a good anti-hemolysis interference performance on BS-2800M system and demonstrated good performances in other assay characteristics. It is therefore suitable for use in clinical laboratories.

B-127 The Effect of Hemolysis on Potassium Measurement in a Community Setting: The Marching Error

Abstract Background Potassium is one of the most sensitive analytes to in-vitro hemolysis. To detect this error, modern clinical chemistry analyzers calculate the hemolysis index (HI) to estimate the degree of specimen hemolysis. Recently, Roche Diagnostics lowered the acceptable potassium HI limit from 100 to 20 and recommended laboratories avoid using specimens with an HI&amp;gt;20. The goal of this study was to determine a clinically acceptable HI cut-off for potassium at our community reference laboratory, relying on patient data from routine laboratory operation. Methods Plasma venous potassium, sodium, and HI test results from community patients measured on Roche cobas 8000 ISE and c701 modules between September 2022 to December 2023 were extracted from the Roche Infinity Middleware (n=1,101,897). Data was divided into groups based on HI which ranged from 0 to 300. The sample pool with the lowest level of hemolysis (HI≤10) was defined as the base group. Mean and standard deviation of potassium for each HI group was calculated and the difference in the mean between each group compared to the base group was calculated as both absolute and percent ΔK. Percent changes in critical values for both potassium and sodium as a function of hemolysis were calculated for each HI group. Measurement uncertainty (MU) for potassium was calculated using a top-down approach. Data were analyzed in Rstudio and Microsoft Excel. Results The vast majority of potassium specimens (83.8%) were stratified into the HI base group (HI ≤10) and 96.8 % had an HI ≤ 20, the cut-off recommended by Roche. The mean potassium level for each HI group increased as the degree of hemolysis in the specimens became more elevated. A MU-derived analytically significant shift of ΔK&amp;gt;0.1 mmol/L was observed for the bin HI 21-30. Depending on the employed total allowable error limit, clinically significant shifts in mean potassium levels were observed at HI bin 41-50 (RCPA: ΔK&amp;gt;0.2 mmol/L) or HI bin 101-150 (EFLM-derived reference change value: ΔK&amp;gt;10.1% and CLIA: ΔK&amp;gt;0.5mmol/L. The shift in patient potassium values seen as HI increased led to a change in critical results reporting frequencies. In the base group (HI≤10), 0.06% and 0.07% of results had low (&amp;lt;2.6 mmol/L) or high (&amp;gt;6.2 mmol/L) critical potassium values, respectively. When HI was 101-300, 0.00 and 15.7% of results had low or high critical potassium values. A similar comparison performed with sodium did not show the same shift in reported critical values. Linear regression of ΔK plotted vs HI produced the equation ΔK=0.0051xHI, validating the equation of Martinez-Morillo and Alvarez published previously using paired hemolyzed/non-hemolyzed potassium laboratory data. Conclusions In community patients, in-vitro hemolysis produced analytically significant errors at an HI of 20. Clinically significant errors occurred between an HI of 40-100. Past HI 100, mean ΔKs of 0.5-1.20 mmol/L were seen in our population. This shift in patient values as a function of hemolysis led to a change in critical result reporting, suggesting the generation of both factious critically high potassium results and the masking of critically low potassium results in our patient population.

B-271 High Sensitivity Homogeneous Enzyme Immunoassay for Benzodiazepines and Metabolites

Abstract Background Benzodiazepines are widely prescribed for treating various conditions such as anxiety, insomnia, and seizures due to their sedative and anxiolytic properties. Despite their therapeutic benefits, concerns persist regarding their potential for misuse, dependence, and addiction. Urine drug screening immunoassays serve as an important tool in monitoring benzodiazepine usage, ensuring adherence to prescribed regimens, identifying misuse or abuse, and facilitating necessary medical interventions. However, limitations of benzodiazepine immunoassays are well-documented in literature, with issues surrounding their lack of sensitivity to glucuronidated and 7-amino metabolites, inability to detect newer benzodiazepines, and poor cross-reactivity with different benzodiazepines. ARK Diagnostics has developed a highly sensitive homogeneous enzyme immunoassay capable of detecting both classic and designer benzodiazepines, in addition to their glucuronide and 7-amino metabolites in human urine at a cutoff concentration 200 ng/mL, without the need for glucuronidase or sample pretreatment. Methods The ARKTM Benzodiazepine Plus Assay is a liquid-stable homogenous enzyme immunoassay consisting of two reagents. The assay uses temazepam as the 200 ng/mL cutoff calibrator. The performance characteristics of this assay, including precision, spiked recovery, specificity, and method comparison to LC-MS/MS, were evaluated on the Beckman Coulter AU680 automated clinical analyzer. Results The ARKTM Benzodiazepine Plus demonstrated acceptable precision, with no overlap between cutoff (200 ng/mL) and ±25% control levels (150 ng/mL and 250 ng/mL) in a histogram overlap analysis, and ≤3.6% CV in semi-quantitative mode. Spiked temazepam samples spanning the semi-quantitative assay range of 1000 ng/mL were recovered between 95.0% and 103.7% of the spiked levels. Thirty-eight benzodiazepines produced cross-reactivities &amp;gt;80%, including the following benzodiazepines and metabolites: lorazepam, clonazepam, diazepam, alprazolam, temazepam, oxazepam glucuronide, lorazepam glucuronide, temazepam glucuronide, 7-aminoclonazepam, and 7-aminoflunitrazepam. In method comparison studies, a total of 103 urine samples containing benzodiazepines with LC-MS/MS values ranging from 1.3 ng/mL to 3257.6 ng/mL were tested with the ARKTM Benzodiazepine Plus Assay. Eighty samples tested positive with values ≥200 ng/mL and 23 samples tested negative with values &amp;lt;200 ng/mL by the ARKTM Benzodiazepine Plus assay. Of the 23 samples that tested negative, 21 had LC-MS/MS values &amp;lt;200 ng/mL and 2 samples were identified as midazolam samples with metabolite a-OH-midazolam levels &amp;gt;200 ng/mL. Conclusions The ARKTM Benzodiazepine Plus Assay enables the sensitive, rapid, and reliable measurement of benzodiazepines and their metabolites in human urine, without the need for hydrolysis or pretreatment. The assay addresses the present constraints of currently available benzodiazepine immunoassays, including their insufficient sensitivity, limited cross-reactivity to metabolites, and the requirement for glucuronidase pretreatment. The ARKTM Benzodiazepine Plus Assay is applicable to a wide range of clinical chemistry analyzers.

Biodistribution and toxic potential of silver nanoparticles when introduced to the female rat reproductive tract

The prevalence of ionic silver and silver nanomaterials in hygiene products has been increasing due to their antimicrobial activity. While numerous studies have examined the effects of nanosilver in laboratory settings, there is a limited understanding of its impact on reproductive tissues, as well as its biodistribution and toxicity upon intra-vaginal exposure. If ionic or nanosilver enters adjacent and internal tissues via intra-vaginal exposure, the overuse of hygiene products containing silver may potentially threaten woman's health. This study investigated the effects of intra-vaginal silver exposure in Female Fischer 344 rats to single and multiple doses of a commercial product containing silver, along with standard nanosilver materials. Custom tampons were developed to simulate practical usage scenarios. The analysis of tissue biodistribution revealed that epithelial penetration and redistribution of silver was observed with most administered silver eliminated in feces (8–44 %), and secondary tissues containing 1–18 % of the dose, predominantly localized in the reproductive tract. In a subsequent toxicity study, vaginal histopathology indicated a cellular inflammatory reaction (neutrophil infiltration) associated with the presence of foreign silver material upon a single administration. Interestingly, no noticeable difference in histopathology incidence was observed upon multiple exposures to silver compared to the control group. Clinical chemistry and hematology analyses following acute exposure to silver nanomaterials showed no significant abnormalities. Overall, acute vaginal exposure to silver nanomaterials and ionic silver resulted in limited silver persistence, local tissue reactivity, epithelial penetration of silver resulting in accumulation in distant organs, and elimination primarily through feces. In vitro data suggested potential alterations in normal vaginal flora. Long-term studies are still lacking in this area.