Laboratory Instruments Research Articles

Differentiation of Acquired Hemophilia a from Lupus Anticoagulant Using Global Coagulation Assays and Magnesium-Dependent Activated Partial Thromboplastin Time Assay

CONCLUSION Rapid and reliable differentiation between bleeding and thrombotic tendency in cases with prolonged activated partial thromboplastin time (APTT) remains challenging in clinical practice, in particular, distinguishing cases positive for coagulation factor inhibitor or lupus anticoagulant (LA). To differentiate acquired hemophilia A (AHA) from LA using three global coagulation assays: rotational thromboelastometry (ROTEM), thrombin generation assay (TGA), and clot waveform analysis (CWA); also to validate the effectiveness of magnesium-dependent APTT (APTT-Mg) assay. This method uses APTT-Mg and normal APTT using Ca to screen for LA, and APTT-Mg/APTT ratio is smaller in LA. We included patients with prolonged APTT without congenital coagulation factor deficiency who were referred to our department between November 2019 and September 2021. APTT prolongation was defined as >35 s (normal range: 25-35 s) measured routinely with Coagpia APTT-N (Sekisui Medical). The AHA group had a confirmed diagnosis and underwent treatment of AHA. The LA group was divided into LA1 (possible LA) and LA2 (definite LA). LA1 was defined by any of the following conditions: (1) index of circulation anticoagulant ≥15 in the cross-mixing test performed with Coagpia APTT-N; (2) positive dilute Russell's viper venom time (DRVVT) phospholipid neutralization method using Gladipore LA (Medical & Biological Laboratories); (3) positive APTT phospholipid neutralization test using Hemosil SCT (Instrumentation Laboratory); (4) positive for anti-CL antibody; (5) positive for anti-CLβ2GPI antibody; or (6) positive for phosphatidylserine-dependent anti-PT antibody. LA2 was defined by index of circulation anticoagulant >12.4 in the cross-mixing test performed by Thrombocheck APTT-SLA (Sysmex Corporation), or item (2) or (3) above, in accordance with criteria of LA in the International Society of Thrombosis and Haemostasis. LA1 and LA2 did not overlap, because cases that did not meet the conditions of LA2 were treated as LA1, and global coagulation assays were performed in all LA cases. In the global coagulation assays, 12 healthy volunteer samples were tested and regarded as the normal goroup. For the APTT-Mg assay, equal volumes of 25 mmol MgCl 2 and 25 mmol CaCl 2 were used instead of the normally used CaCl 2, and Thrombocheck APTT-SLA was used to measure APTT-Mg, which was then divided by the normal APTT concentration measured with the same APTT reagent. Ten patients in the AHA, 12 in the LA1, and 32 in the LA2 groups were eligible. ROTEM showed that clotting time in the non-activated rotational thromboelastometry assay (NATEM) mode was >3600 s (minimum 1363 s) for AHA, 501 s (IQR: 430-632 s) for LA1, 533 s (IQR: 443-641 s) for LA2, and 500 s (IQR: 429-645 s) for the normal group. TGA showed that peak thrombin concentration was 16 nM (IQR: 13-29 nM) in the AHA group, 242 nM (IQR: 175-273 nM) in the LA1 group, 174 nM (IQR: 129-259 nM) in the LA2 group, and 190 nM (IQR: 125-288 nM) in the normal group. In the CWA, Ad|min1| was 2.1 (IQR: 1.7-3.2) in the AHA group, 8.7 (IQR: 8.0-9.5) in the LA1 group, 6.7 (IQR: 5.8-8.4) in the LA2 group, and 10.3 (IQR: 9.8-10.6) in the normal group. APTT-Mg/APTT was 1.23 (IQR: 1.18-1.29), 1.26 (IQR: 1.23-1.38), and 1.05 (IQR: 0.95-1.20) in the AHA, LA1, and LA2 groups, respectively. ROTEM, TGA, and CWA effectively differentiated between the LA1/2 and AHA groups. Clotting time in non-activated rotational thromboelastometry assay mode (ROTEM) and peak height (TGA) were better at distinguishing high-titer LA cases from LA-positive AHA cases. APTT-Mg/APTT could be suitable for distinguishing AHA from high-titer LA; however, it may not be useful for differentiating AHA from low-titer or suspected LA.

Development of Visible Spectrophotometric Methods for the Determination of Tricyclic Antidepressants Based on Formation of Molecular Complexes with p-Benzoquinones

Tricyclic antidepressants are commonly employed in the management of major depressive disorders. The present work describes two visible (VIS) spectrophotometric techniques that utilize the formation of charge transfer complexes between four antidepressant compounds, namely, amitriptyline hydrochloride (AMI), imipramine hydrochloride (IMI), clomipramine hydrochloride (CLO), and trimipramine maleate (TRI) acting as electron donors and two p-benzoquinones, namely, p-chloranilic acid (pCA) and 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ), serving as electron acceptors. The stoichiometry of the compounds produced exhibited a consistent 1:1 ratio in all instances, as established by Job’s method. Molar absorptivities, equilibrium association constants, and several other spectroscopic properties were determined for all complexes. The developed spectrophotometric techniques were validated intra-laboratory and successfully applied for quantitative assessment of the four antidepressant active ingredients in several commercial pharmaceutical formulations. The methods are relatively simple, fast, and use readily available laboratory instrumentation, making them easily applicable by most quality control laboratories worldwide.

Microgravity as an Anti-Metastatic Agent in an In Vitro Glioma Model

Gravity is a primary physical force that has a profound influence on the stability of the cell cytoskeleton. In our research, we investigated the influence of microgravity on altering the cytoskeletal pathways of glioblastoma cells. The highly infiltrative behavior of glioblastoma is supported by cytoskeletal dynamics and surface proteins that allow glioblastoma cells to avoid stable connections with the tissue environment and other cells. Glioblastoma cell line C6 was exposed to a microgravity environment for 24, 48, and 72 h by 3D-RPM, a laboratory instrument recognized to reproduce the effect of microgravity in cell cultures. The immunofluorescence for GFAP, vinculin, and Connexin-43 was investigated as signals related to cytoskeleton dynamics. The polymerization of GFAP and the expression of focal contact structured by vinculin were found to be altered, especially after 48 and 72 h of microgravity. Connexin-43, involved in several intracellular pathways that critically promote cell motility and invasion of glioma cells, was found to be largely reduced following microgravity exposure. In conclusion, microgravity, by reducing the expression of Connexin-43, alters the architecture of specific cytoskeletal elements such as GFAP and increases the focal contact, which can induce a reduction in glioma cell mobility, thereby inhibiting their aggressive metastatic behavior.

Improved Separation in Horizontal Protein SDS-PAGE with Double-Deck Flat Electrodes and a Field Inversion Gel Electrophoresis Module.

The horizontal flatbed electrophoresis method is employed to separate protein samples, providing greater flexibility for various electrophoretic applications and easier sample loading compared to its vertical counterpart. In the currently available equipment setup, cathode and anode electrodes are positioned on top of a gel at each end. Since an electric field enters the gel from the top, its strength gradually weakens from the top to the bottom of the gel. When examining the interior of gels following electrophoretic separation, the uneven electric field causes the protein bands to lie down forward in the direction of migration, leading to an increase in bandwidth. This issue has remained unaddressed for several decades. To address this problem, new clamp-shaped and double-deck electrodes were developed to apply an electric field simultaneously from both the top and bottom of the gel. Both of these new electrodes facilitated the formation of perpendicular protein band shapes and enhanced resolution at a comparable level. Due to their ease of use, double-deck electrodes are recommended. By combining these new electrodes with the field inversion gel electrophoresis (FIGE) technique, the protein bands could be focused and aligned nearly vertically, resulting in the highest level of electrophoretic resolution. Our electrodes are compatible with polyacrylamide gels of varying sizes, buffer systems, and sample well formats. They can be easily manufactured and seamlessly integrated into existing laboratory instruments for practical use.

Comprehensive Characterization of Coagulation Parameters in Venous Malformations

Introduction: Localized intravascular coagulopathy (LIC) is a well-recognized, though poorly characterized complication of venous malformations (VM) that can lead to bleeding, thrombosis, and phlebolith formation. While LIC has classically been characterized by elevations in D-dimer and reductions in fibrinogen, no comprehensive studies of coagulation parameters in VM have been performed to date. Since 2021, all patients with VM undergoing evaluation for LIC in the Yale Vascular Malformations Program (VaMP) and the Yale Classical Hematology clinic have been subjected to an extensive set of coagulation tests to fully analyze LIC. Our aim was to use these laboratory test results to comprehensively characterize LIC in this population. Methods: We conducted a retrospective chart review of all VM patients presenting to the Yale VaMP and the Yale Classical Hematology clinic for assessment of LIC from 2021 to 2023. All included patients were evaluated for LIC using the following coagulation parameters: von Willebrand Factor (VWF) antigen, VWF activity, factor VIII (FVIII), alpha-2 antiplasmin (A2AP), plasminogen activator inhibitor-1 (PAI-1), thrombin-antithrombin complex (TAT), D-dimer (DD), fibrinogen, prothrombin time (PT), international normalized ratio (INR), and partial thromboplastin time (PTT). Measurements of VWF antigen, VWF activity, FVIII and A2AP were performed using an ACL TOP system (Instrumentation Laboratory; Bedford, MA, USA), while D-dimer and fibrinogen were measured using a BCS XP System (Siemens; Malvern, PA, USA) at our institution's clinical laboratory. PAI-1 and TAT were processed at national Clinical Laboratory Improvement Amendment-certified reference laboratories using ELISA-based assays. Baseline patient characteristics and coagulation test results were extracted via manual chart review. Data analysis was done using IBM SPSS statistics software and GraphPad Prism 9 software. Categorical variables were described using frequency and percentages, while quantitative variables were described using central tendency and dispersion measures. We performed univariate analysis using the Chi-square test. We also analyzed the coagulation tests using a correlation matrix. Statistical significance was set at p<0.05. This project was approved by our Institutional Review Board. Results: A total of 23 patients with VM were included in the analysis (Table 1). The mean age was 38 ± 16 years; the majority (82.6%) were female. The most common anatomic location was the head and neck (52.2%), while the most frequent extent of tissue involvement was muscle (54.5%), with most patients having a single lesion (59.1%) rather than multiple ones. No patients had venous thromboembolism. Normal DD and high TAT levels were observed in most patients (68.2% and 69.6%, respectively). VWF activity, VWF antigen, FVIII, PAI-1, and fibrinogen all were positively correlated (Figure 1). TAT was positively correlated with PAI-1 and inverselycorrelated withPT and INR. DD was positively correlated only with vWF activity. TAT and DD had a poor correlation; among patients with a normal TAT (n=7), 85.7% had a normal DD, while among patients with a normal DD (n=15), only 40% had a normal TAT. Three out of 4 (75%) patients with skin involvement had a high TAT with normal DD, while 2 out of 3 (66.7%) patients with visceral involvement had both high TAT and DD, although these patternswere not statistically significant (P=0.42 and 0.31, respectively). Among patients with muscle involvement (n=11), 45.5% had a high TAT and normal DD while 36.3% had both high TAT and DD. Conclusions: In this first comprehensive hematologic study of VM, we demonstrate that VM-related LIC is characterized by derangements of multiple coagulation parameters. Due to this, measurements of DD and fibrinogen alone may be inadequate for assessing LIC, and the addition of TAT and multiple other coagulation tests may yield a more complete picture of hemostatic derangements in LIC. A lack of correlation between DD and TAT is unexpected and merits further study. Differences in the depth of the VM tissue involvement and TAT/DD correlation may indicate a progression of coagulopathy related to the extent of VM but require further investigation in a larger study.

Radchip: A Laminin-Lined Red Blood Cell (RBC) Adhesion Device for Functional Characterization and Clinical Evaluation of Sickle RBCs at Steady-State

Sickle cell disease (SCD) is a genetic disorder caused by a point mutation in the beta-globin gene producing an abnormal hemoglobin (HbS) which, under deoxygenation, polymerizes and causes severe distortion of the red blood cell (RBC). RBCs may be dehydrated, poorly deformable and highly adhesive to the endothelium, with abnormal adhesion being of primary interest to this study. Recently, several new disease modifying drugs have been FDA approved, with more in development. These novel agents target specific aspects of SCD pathophysiology, including adhesion. Changes induced by such therapies are not always detectable by common conventional lab instruments. LORRCA (oxygen-gradient ektacytometry) is the gold-standard for deformability measurement; it has been well described both analytically and clinically in SCD. Adhesion and deformability have a dynamic, yet complementary relationship that we explore here. While these may behave in an interdependent manner, each determinant must be considered independently. There is a growing demand for innovative tools to evaluate rheological properties under physiological conditions, particularly microfluidic devices. However, many current devices are limited in their clinical utility due to, choice of ligand, complexity of design, or reliance on endothelial cell culture (typically HUVEC cells) which are difficult to use, vary with passage number, and do not represent adult SCD endothelium. We developed a Red blood cell Adhesion Chip (RAdChip) to model RBC to endothelial adhesion using laminin, a physiologically relevant ligand, to easily, effectively, and reproducibly measure RBC adhesion. Here, we describe our analytic and clinical validation of our device. Peripheral blood samples were collected in EDTA from individuals with SCD and healthy controls under an IRB approved protocol. RBCs were extracted via centrifugation, washed and resuspended in PBS to a 0.5% HCT. Microfluidic devices were fabricated using polydimethylsiloxane (PDMS). A 5ug/ml solution of recombinant laminin was applied to each channel, then blocked with 1% bovine serum albumin (BSA). Resuspended RBCs were introduced to the device through tubing connected to the inlet port. Harvard Apparatus syringe pumps were used to perfuse samples and wash channels at ~1dyne/cm 2. Adhesion was visualized with a Keyence inverted bright-field microscope at 20x (Itasca, IL). Adhered cells were counted within a 1.8mm 2 region of interest (Roi) using the iCLOTS automated cell quantification software (Fay et al, Nat comm accepted 7/2023). We measured adhesion index via RAdChip for common SCD genotypes: HbSS/SB0: (median (x͂) =756; range (r): 154-1889; n=26) HbSC/SB+ : (x͂ =299; r=: 114-731; n=11 ) HbAA: (x͂) =13; r= 5-90; n=7). RBC adhesion values were independent of age (p=0.65) and gender (p=0.63). An increase of 0.1 unit in EImin and EImax decreases RBC adhesion by 214 and 268 cells/mm 2, respectively. A 1mmHg increase in PoS increased adhesion by 19.5 cells/mm 2. sRBC adhesion had a significant positive association with vaso-occlusive events (VOE) (p=0.005), absolute neutrophil count (ANC) (p=0.005), and EImax (p<0.001). Notably, analytical validation of RAdChip produced a mean CV of 0.13 ± 0.94. RAdChip adds a key component of functional RBC evaluation that conventional lab equipment does not measure. We have analytically validated our device, demonstrating an excellent coefficient of variability. 44 patient samples, each examined in triplicate produced a mean CV of 0.13. We have demonstrated clinical validity of our device by establishing correlations with known markers of severity and pain event frequency. RBC adhesion values increase as the number of VOE events increase, reaffirming the belief that adhesion is a principal element in the multistep VOE cascade. In like manner, improvements in deformability are associated with decreased RBC adhesion, strengthening the interdependent nature of VOE molecular events. Recognizing the dynamic nature of SCD, it is necessary to explore sickle erythrocyte adhesion longitudinally to investigate changes before, during, and after VOE occurrences. Continued and increased integration of adhesion measurements with RAdChip and other rheology devices into routine SCD care may be helpful in assessing disease severity, identifying need for additional disease modifying agents, and assist with second line agent selection and monitoring.

A Tunable Dielectric Resonator Oscillator with Phase-Locked Loop Stabilization for THz Time Domain Spectroscopy Systems

Precision microwave measurement systems require highly stable oscillators with both excellent long-term and short-term stability. Compared to components used in laboratory instruments, dielectric resonator oscillators (DRO) offer low phase noise with greatly reduced mechanical complexity. To further enhance performance, phase-locked loop (PLL) stabilization can be used to eliminate drift and provide precise frequency control. In this work, the design of a low-cost DRO concept is presented and its performance is evaluated through simulations and measurements. An open-loop phase noise of −107.2 dBc/Hz at 10 kHz offset frequency and 12.8 GHz output frequency is demonstrated. Drift and phase noise are reduced by a PLL, so that a very low jitter of under 29.6 fs is achieved over the entire operating bandwidth.

Operationalisation of the ForenSeq® Kintelligence Kit for Australian unidentified and missing persons casework

Single nucleotide polymorphism (SNP) genotyping technologies can generate investigative leads for human remains identification, including estimation of biological sex, biogeographical ancestry (BGA), externally visible characteristics (EVCs), identity, uniparental lineage and extended kinship. The ForenSeq® Kintelligence Kit provides forensic laboratories with the ability to apply this suite of genetic tools to forensic samples using one panel targeting 10,230 SNPs (including 56 ancestry-informative, 24 phenotype-informative, 94 identity-informative, 106 X chromosome, 85 Y chromosome and 9867 kinship-informative SNPs) sequenced on the MiSeq FGx® Sequencing System. The ForenSeq® Kintelligence Kit has been internally validated, optimised and operationalised by the Australian Federal Police National DNA Program for Unidentified and Missing Persons (AFP Program) for coronial casework. The internal validation was conducted according to the Scientific Working Group on DNA Analysis Methods guidelines (excluding mixture analysis), focussing on sample types typically encountered in human remains identification casework, such as bones, teeth, nail, blood and hair. The workflow was optimised for a high throughput library preparation and sequencing workflow, and additional analytical thresholds were developed to improve genotyping accuracy for low DNA input samples. Additionally, the genetic intelligence generated from the kit was compared to the self-declared biological sex, EVCs and BGA of the DNA donors to assess concordance. The kit was able to produce high quality SNP profiles from 1.0 ng down to 0.1 ng of DNA, with high repeatability and reproducibility, and minimal background noise. The prediction accuracy for biological sex (95%), hair colour (58%), eye colour (74%) and BGA inferences (consistent: 74%; partially consistent: 10%; inconclusive: 16%) was determined based on self-declared data. Additionally, SNP profiles from a volunteer family group of ten related individuals were uploaded to GEDmatch PRO™ to assess kinship accuracy. The kit was capable of detecting (97%) and accurately classifying (90%) genetic relationships spanning from first to fifth degree. The Kintelligence Kit provides the AFP Program with a robust and reliable genetic intelligence tool for unidentified and missing persons investigations, which has been designed to sequence multiple challenging samples in a single multiplexed assay using existing laboratory instrumentation.

FACE-TO-FACE VS. ONLINE LEARNING in SCIENCE and ENGINEERING COURSES DURING COVID-19 EPOCH

ABSTRACT Background The new teaching scenario after COVID-19 results in the adoption of a complete online environment, providing a unique opportunity to compare in site (face-to-face) vs virtual (online) teaching. Due to the pandemic, practical activities have been replaced by written protocols supported with photos, video-recorded demos, linearsimulations, remote-controlled real laboratory instruments and so forth. Purpose This article describes the adaptation from in site to online teaching in science and engineering subjects in a Spanish University during COVID-19. Advantages and drawbacks, along with the process adopted (from education tools through virtualization tools) are comprehensively explained, including an inquiry-based learning (IBL) methodology for teaching laboratory courses. Sample Courses in Chemistry (cohort size of 100 students), Pharmacy (50 students), Industrial Electronics and Automation Engineering (110 and 30 students) at the University of Alcalá. Methods The outcomes of the teaching-learning process were evaluated via online tests and rubrics. The impact of online adaptation was assessed by comparing the marks of the different test and the dropout rate in the subjects with those obtained from groups of previous years in the face-to-face mode and feedback from the students. Anonymous survey was conducted to attain a comprehensive understanding of the students’ experiences with virtual learning. Results The developed approaches are beneficial for the teaching-learning process and lead to a higher number of students passing the subjects compared to the face-to-face modality. Some challenges students encountered were internet connection, managing workload andovercoming distractions. Nonetheless, most students benefited from the flexibility and better accessibility to the course material, timesaving and relax environment. Conclusion Our findings provided evidence in favor of remote learning. Both students and instructors regard the strategy as very positive. The new methodologies reported herein can be extended to online teaching courses beyond the COVID-19 scenario.

From PID to Set Point Weighted PID: A Teaching DC-DC Buck Converter Platform

This article presents a lowcost educational platform based on a DC-DC Buck Converter to implement real-time control laws for engineering students. The motivation for this work arises from what happened during the COVID-19 pandemic when students did not have access to laboratory equipment and instruments necessary for their academic development. Particularly in the subjects of electrical circuits, electronics, and control, there has been a significant drop in the development of projects. This work shows the Buck converter's modeling and design to build and program different control laws like the Proportional Integral Derivative and the Setpoint weighted PID. Real-time experiments show the performance of the DC-DC Buck converter. The control algorithms are implemented using the MatLab / Simulink programming platform under the Waijung real-time control environment and an STM32704 data acquisition card.

Application of a diagnostic algorithm for differential diagnosis in a patient with recurrent abdominal pain

Introduction Among the diseases of urgent pathology the leading place belongs to acute pancreatitis, which is second only to acute appendicitis and acute cholecystitis. Moreover, a significant share is occupied by mild forms, which account for 85%. In the Clinical Recommendations for Acute pancreatitis in the section “Prevention and dispensary monitoring”, the authors recommend regular dispensary monitoring by a gastroenterologist and a surgeon. However, this nosology according to the Order of the Ministry of Health of the Russian Federation from 03.15.2022 No 168N is not included in the dispensary observation. In this regard, patients remain without proper attention of doctors when they are discharged from hospitals, or when they refuse hospitalization after examination in the emergency room.The aim of the work – is to present the result of applying the diagnostic algorithm of differential diagnosis of diseases and detection of gastrointestinal tract pathology on the clinical example of a patient with recurrent abdominal pain syndrome.Materials and methods The analysis of medical records of a 59-year-old patient who repeatedly applied to the network of polyclinics and hospitals providing emergency specialized medical care for recurrent abdominal syndrome was carried out.Results and discussion In this clinical observation of recurrent abdominal syndrome in a patient, the biliary mask of acute pancreatitis was revealed. In outpatient conditions using the developed diagnostic algorithm, choledocholithiasis, juxtapapillary diverticulum of the duodenum and hernia of the esophageal aperture of the diaphragm were detected. The diagnosis made it possible to perform the treatment competently and lead to the patient's recovery. Thanks to the dispensary observation and additional methods of investigation according to the developed algorithm, the following were detected: juxtapapillary diverticulum of the duodenum, which may not be visualized at video esophagogastroduodenoscopy with direct optics, as well as ductal stones, which are not always visualized at transabdominal ultrasound and are detected at endoscopic retrograde cholangiopancreatography or endoscopic ultrasonography. Difficulties may arise in the diagnosis of gastrointestinal diseases due to the variability of the clinical course and polymorphism of symptoms in this category of patients. Such a course of diseases causes difficulties at the stage of patients' admission to the emergency room of clinics, can lead to diagnostic errors that lead to incorrect treatment tactics.Conclusion The application of an algorithm based on the assessment of clinical criteria in conjunction with instrumental imaging methods and laboratory tests allowed to optimize the diagnostic process and facilitated the choice of optimal therapy and surgical tactics for the treatment of the patient with recurrent abdominal pain syndrome.

A novel serum calprotectin (MRP8/14) particle-enhanced immuno-turbidimetric assay (sCAL turbo) helps to differentiate systemic juvenile idiopathic arthritis from other diseases in routine clinical laboratory settings

BackgroundDifferential diagnosis in children with signs of unprovoked inflammation can be challenging. In particular, differentiating systemic juvenile idiopathic arthritis (SJIA) from other diagnoses is difficult. We have recently validated the complex of myeloid-related proteins 8/14 (MRP8/14, also known as S100A8/A9 complex or serum calprotectin) as a helpful biomarker supporting the diagnosis of SJIA. The results were subsequently confirmed with a commercial ELISA. However, further optimization of the analytical technology is important to ensure its feasibility for large-scale use in routine laboratory settings.MethodsTo evaluate the accuracy in identifying children with SJIA, the performance of a particle-enhanced immuno-turbidimetric assay for serum calprotectin (sCAL turbo) on an automated laboratory instrument was analyzed. Samples from 615 children were available with the diagnoses SJIA (n = 99), non-systemic JIA (n = 169), infections (n = 51), other inflammatory diseases (n = 126), and acute lymphoblastic leukemia (ALL, n = 147). In addition, samples from 23 healthy controls were included.ResultsThe sCAL turbo assay correlated well with the MRP8/14 ELISA used in previous validation studies (r = 0.99, p < 0.001). It could reliably differentiate SJIA from all other diagnoses with significant accuracy (cutoff at 10,500 ng/ml, sensitivity 84%, specificity 94%, ROC area under curve 0.960, p < 0.001).ConclusionsSerum calprotectin analyses are a helpful tool supporting the diagnosis of SJIA in children with prolonged fever or inflammatory disease. Here, we show that an immuno-turbidimetric assay for detection of serum calprotectin on an automated laboratory instrument can be implemented in clinical laboratory settings to facilitate its use as a diagnostic routine test in clinical practice.

Implementation QR-Code for Introducing Optical Instruments

The laboratory is one of the important facilities and supports the implementation of learning in tertiary institutions. However, in carrying out practicum activities, students are often constrained by information regarding names, functions, and steps for using instruments. Therefore, the need for information about laboratory equipment is quite important for students. This study aims to produce a website based on a QR code to provide information about the instrument to users and the steps to use it. This research uses research and development methods with the ADDIE model (Analysis, Design, Develop, Implementation, and Evaluation). The questionnaire is used to measure user responses regarding the QR code and its application. Based on the analysis of the problems found, the QR code was successfully created. This QR code was validated by media experts and instrument content experts and obtained valid results. Users show a positive response regarding the use of the QR code for introducing of Refraction Laboratory instruments. Thus, a website-based QR code can be used to provide optical instrument information to users. However, socialization about its use is very necessary for laboratory users.

Open Hardware for Microfluidics: Exploiting Raspberry Pi Singleboard Computer and Camera Systems for Customisable Laboratory Instrumentation.

The integration of Raspberry Pi miniature computer systems with microfluidics has revolutionised the development of low-cost and customizable analytical systems in life science laboratories. This review explores the applications of Raspberry Pi in microfluidics, with a focus on imaging, including microscopy and automated image capture. By leveraging the low cost, flexibility and accessibility of Raspberry Pi components, high-resolution imaging and analysis have been achieved in direct mammalian and bacterial cellular imaging and a plethora of image-based biochemical and molecular assays, from immunoassays, through microbial growth, to nucleic acid methods such as real-time-qPCR. The control of image capture permitted by Raspberry Pi hardware can also be combined with onboard image analysis. Open-source hardware offers an opportunity to develop complex laboratory instrumentation systems at a fraction of the cost of commercial equipment and, importantly, offers an opportunity for complete customisation to meet the users' needs. However, these benefits come with a trade-off: challenges remain for those wishing to incorporate open-source hardware equipment in their own work, including requirements for construction and operator skill, the need for good documentation and the availability of rapid prototyping such as 3D printing plus other components. These advances in open-source hardware have the potential to improve the efficiency, accessibility, and cost-effectiveness of microfluidic-based experiments and applications.

Wearable and recyclable epinephrine biosensors based on molecular imprinting polymer modified organic electrochemical transistors

Tracking fluctuations of epinephrine level is essential for doctors or patients to evaluate the body’s condition, but traditional epinephrine detection completely relies on the laboratory instrument. Herein, a wearable and recyclable epinephrine sensor based on the organic electrochemical transistor (OECT) has been developed for rapid, simple, and continuous detection. The molecular imprinting polymer (MIP) is modified on the gate electrode which can recognize the epinephrine molecules, and the voltage drop caused by redox of epinephrine will generate sensing signal. For epinephrine molecules, the boronate affinity binding in MIP and oxidation on electrodes make the sensor have great recyclability. The detection limit of this sensor is 10 pM, which is suitable for humoral metabolites. The sensor can be used for detection at least 20 cycles and recovered by the simple positive voltage cleaning, which is economical for wearable device. The flexible sensors units based on the polydimethylsiloxane (PDMS) are constructed with a matching portable circuit system connecting to the computer and mobile phone APPs. The whole wearable sensor puts up a user-friendly detection method with good recyclability, stability, selectivity and low detection limit for doctors or patients.

Development of an optical particle number sensor based on continuous wave laser-induced incandescence

In this work, the development of a new particle counting sensor based on continuous-wave laser-induced incandescence is described. We demonstrate the sensor, which makes use of a focussed laser beam emitted by a compact laser diode, as an alternative device to established laboratory techniques to count soot particles. Simulations help understanding the cause–effect relationships based on known theory with the implementation of specific laser beam characteristics that influence the absorption process. The main goal of this work was to demonstrate the sensor’s ability to determine the soot particle number concentration of a test aerosol. This has been confirmed through a comparison with established laboratory instruments, like scanning mobility particle sizer or condensation particle counter. The observed size dependency of the laser induced incandescence sensor’s detection efficiency was successfully corrected with prior knowledge of the investigated particle size distribution. Furthermore, the sensor’s detection limit with respect to the particle diameter was determined to be well below 100 nm with a dependency on the applied laser power.

Experimental investigation into the effects of voids on the response of buried flexible pipes subjected to incrementally increasing cyclic loading

In this study, large-scale fully instrumented laboratory tests were conducted to investigate the behaviour of buried flexible high-density polyethylene (HDPE) pipes, in sand beds with and without voids subjected to incrementally increasing cyclic loading. Voids with a predetermined size were created at one side of the spring-lines of the pipes, which were buried at variable depths, H, of 1.5, 2.0 and 2.5 times the diameter of the pipe, D. Results showed that increasing the pipe burial depth, H/D, contributed to decreasing the settlement of the footing, deformation of the pipe crown and invert, lateral displacement of the spring-line, and the stress and strain generated along the pipe crown and invert. Void presence led to a significant increase in the footing settlement, which ranged from 3 % up to 18 %, according to H/D. Furthermore, void presence led to a sharp increase in the crown, invert, and spring-line settlements, which ranged from 34 % to 52 %, 10 %–12.5 %, and 13 %–38 %, respectively. Increasing pipe burial depth was found to be highly effective in protecting buried pipes, minimising inevitable consequences of the presence of voids. However, this was combined with an increase in the pressure at the pipe spring-line that led to a positive horizontal support at the pipe's spring-lines resulting in reducing pipe deformation.

Genetic Polymorphism of Lipoprotein-Associated Phospholipase A2 Influences Susceptibility to Gestational Diabetes Mellitus in Chinese Population.

This paper aims to study the relationship between lipoprotein-associated phospholipase A2 (Lp-PLA2) and GDM (gestational diabetes mellitus) by detecting Lp-PLA2 level and its gene polymorphism. From January to June 2022, 82 GDM patients treated in our hospital were included as an experimental group, and 89 healthy pregnant women during the same period were selected as the control group. Lp-PLA2 concentration and TG, TC, HDL-C, and LDL-C levels were tested with specialized instruments in clinical laboratories. The PLA2G7 gene polymorphisms (rs1805017, rs1805018, and rs76863441) were detected by fluorescent probe method and sequencing. Lp-PLA2 concentration was significantly higher in GDM group than control group (P<0.05). Among three polymorphism loci of PLA2G7 gene (rs1805017, rs1805018, and rs76863441) the significant associations were only found in GT genotype of rs76863441 loci (P<0.05). Pregnant women with high levels of Lp-PLA2 concentration are more likely to develop GDM, especially those with PLA2G7 rs76863441 polymorphism. Lp-PLA2 concentration and PLA2G7 rs1805017 polymorphism may be a novel marker for GDM diagnosis and prediction.

A-140 Evaluating the Performance of Various Lupus Anticoagulants Tests Among Different Coagulation Platforms

Abstract Background Lupus Anticoagulants (LAs), one of the antiphospholipid antibodies, are risk factors for arterial or venous thrombosis with complications such as stroke, acquired thrombophilia, and pregnancy loss. Detection of LAs is challenging for laboratories as the heterogeneity of antibodies and epitopes on protein in the phospholipids. Thus, no single test is sensitive for all LAs, and the International Society on Thrombosis and Haemostasis Scientific and Standardization Committee (ISTH) recommended laboratory should provide two test systems based on different principles, which dilute Russell's viper venom time (dRVVT) should be as the major test, and partnered with LA-sensitive activated partial thromboplastin time (aPTT). There are many different mechanisms of tests proposed by various manufacturers, however, the comparison of different platforms by different manufacturers on LAs remains unknown. In this study, we aimed to evaluate the performance of various LA tests among different platforms. Methods A total of 76 specimens from September to December 2022 were collected. dRVVT (dRVVTIL) and silica clotting time (SCTIL) were performed by ACL TOP 500 analyzer (Instrumentation Laboratory, US). In addition, ellagic acid-based aPTT (Siemens Dade Actin FSL reagent, aPTTAFSL), silica-based aPTT (Siemens Pathromtin SL reagent, aPTTPSL), and dRVVT (dRVVTsysmex) were performed by Sysmex CN-6000 analyzer (Sysmex Corporation, Japan). Analytical performances, agreements, and McNemar's test were calculated. Results All 76 specimens were originally tested by dRVVTIL and SCTIL, and showed that 30 specimens were positive for both assays, 6 were only positive for dRVVT, 5 were only positive for SCT, and 35 specimens were negative for both assays. In addition, all samples were further tested by dRVVTsysmex. Compared with the integrated use of dRVVTIL and SCTIL, dRVVTsysmex showed 100.0% (41/41) positive agreement and 91.4% (32/35) negative agreement. 3 specimens could only be tested positive on the dRVVTsysmex platform, of which one patient was diagnosed with systemic lupus erythematosus (SLE), another patient was diagnosed with infertility, and the other patient was diagnosed with antiphospholipid syndrome (APS). To further evaluate the performance of aPTT on LA screening, 44 specimens with either dRVVTIL, SCTIL and dRVVTsysmex positive were considered LA-positive and further tested by different aPTT assays. The positive agreement was 65.9% (29/44; 95% CI: 50.1–79.5%) and 77.3% (34/44; 95% CI: 62.2–88.5%) on aPTTAFSL and aPTTPSL, respectively, however, there was no significant difference between the positive agreement of two assays (P = 0.13). Conclusion Compared with the integrated use of dRVVTIL and SCTIL, dRVVTsysmex could achieve the same detection rate on LA. Furthermore, aPTTAFSL and aPTTPSL showed no significant difference between the positive agreements for LA-positive specimens.

A-369 Evaluation of a Point-of-Care Whole Blood Creatinine Assay and eGFR Concordance using the CKD-EPI 2021 Equation

Abstract Background Estimated glomerular filtration rate (eGFR) from serum creatinine is considered a better assessment of renal function than serum creatinine alone. Patients with chronic kidney disease have a higher risk of developing contrast-induced nephropathy (CIN). GEM Premier ChemSTAT (Instrumentation Laboratory, Bedford, USA) whole blood (WB) creatinine assay along with eGFR reporting capability per the Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) 2009 eGFR equation is now available and could provide a quick point-of-care (POC) screening test alternative to the lab assay for patient risk management. Currently the CKD-EPI 2009 formula for non-AA population has been used to estimate eGFR in our institution. Recently the CKD-EPI 2021 equation without race adjustment has been recommended by multiple professional societies. The goal of this clinical evaluation was to compare the analytical performance of the ChemSTAT WB creatinine assay and the clinical concordance of eGFR to an established lab method by CKD-EPI 2021 equation. Methods Remnant heparinized WB samples along with age and gender were obtained. The WB samples were analyzed on the GEM Premier ChemSTAT and the plasma assayed on an Architect analyzer (Abbott Laboratories, Lake Forest, USA) as the reference method. The eGFR based on the CKD-EPI 2009 equation were obtained for both WB and plasma creatinine. The eGFR based on CKD-EPI 2021 equation were retrospectively calculated based on WB and plasma creatinine, age and gender. The clinical concordance of categorizing patients with abnormal kidney function (eGFR &amp;lt; 60 ml/min/1.73 m2) and overall concordance of patients (eGFR &amp;lt; 60 + eGFR≥60) was assessed for WB vs plasma eGFR. Error grid analysis was performed for eGFR as described by Snaith et al. (1) to identify the impact of discordant results between WB and plasma eGFRs. Results A total of 60 patient samples were analyzed, median age (SD) = 68.5 (15.2), 53.3% female. The WB ChemSTAT creatinine assay correlated well with the Architect across the tested ranges with regression results of: ChemSTAT_Creatinine (μmol/L) = 1.1099*Architect_Creatinine – 16.0, r = 0.990. A small mean bias (SD) of -3.0 (22.6) μmol/L was observed between ChemSTAT and Architect from the Bland-Altman analysis. When identifying patients with abnormal kidney function (eGFR &amp;lt; 60), ChemSTAT WB eGFR showed 84% (21/25) concordance against the plasma eGFR. An overall clinical eGFR concordance of 88% (53/60) was observed for ChemSTAT. The error grid analysis indicated 100% (60/60) of ChemSTAT WB eGFRs (Zones A and B combined) had no implication to clinical management. Mean biases (SD) from reported CKD-EPI eGFR 2021 vs 2009 equation of 3 (1.6) and 2 (2.6) were observed from Architect plasma creatinine and ChemSTAT WB creatinine, respectively. Conclusions Strong correlations were observed between the ChemSTAT WB POC creatinine assay and eGFR versus the lab method. Based on the high clinical concordance of WB eGFR with CKD-EPI 2021 equation and low mean bias, GEM Premier ChemSTAT demonstrated good performance as POC method for rapid and lab quality renal function assessment.