Sample Tube Research Articles

Impact of sample processing delays on plasma markers of inflammation, chemotaxis, cell death, and blood coagulation.

Biosampling studies in critically ill patients traditionally involve bedside collection of samples followed by local processing (ie. centrifugation, aliquotting, and freezing) and storage. However, community hospitals, which care for the majority of Canadian patients, often lack the infrastructure for local processing and storage of specimens. A potential solution is a "simplified" biosampling protocol whereby blood samples are collected at the bedside and then shipped to a central site for processing and storage. One potential limitation of this approach is that delayed processing may alter sample characteristics. To determine whether delays in blood sample processing affect the stability of cytokines (IL-6, TNF, IL-10, IFN-γ), chemokines (IL-8, IP-10, MCP-1, MCP-4, MIP-1α, MIP-1β), cell-free DNA (cfDNA) (released by dying cells), and blood clotting potential in human blood samples. Venous blood was collected into EDTA and citrate sample tubes and stored at room temperature (RT) or 4°C for progressive intervals up to 72 hours, prior to processing. Plasma cytokines and chemokines were quantified using single or multiplex immunoassays. cfDNA was measured using Picogreen DNA Quantification. Blood clotting potential was measured using a thrombin generation assay. Blood samples were collected from 9 intensive care unit (ICU) patients and 7 healthy volunteers. Admission diagnoses for the ICU patients included sepsis, trauma, ruptured abdominal aortic aneurysm, intracranial hemorrhage, gastrointestinal bleed, and hyperkalemia. After pre-processing delays of up to 72 hours at RT or 4°C, no significant changes were observed in plasma cytokines, chemokines, cfDNA, or thrombin formation. Delayed sample processing for up to 72 hours at either RT or 4°C did not significantly affect cytokines, chemokines, cfDNA, or blood clotting potential in plasma samples from healthy volunteers and ICU patients. A "simplified" biosampling protocol is a feasible solution for conducting biosampling research at hospitals without local processing capacity.

Water vapor stable isotope memory effects of common tubing materials

Abstract. Water molecules in vapor can exchange with gaseous water molecules sticking to surfaces of sampling tubing, and exchange rates are unique for each water isotopologue and tubing material. Therefore, water molecules on tubing walls take some time to reach isotopic equilibrium with a new vapor isotopic signal. This creates a memory effect that is observed as attenuation time for signal propagation in continuous stable water vapor isotope measurement systems. Tubing memory effects in δD and δ18O measurements can limit the ability to observe fast changes, and because δD and δ18O memory are not identical, this introduces transient deuterium excess (D-excess, defined as δD-8×δ18O) artifacts in time-varying observations. To our knowledge, a comprehensive performance comparison of commonly used tubing material water exchange properties in laser-based measurement systems has not been published. We compared how a large isotopic step change propagated through five commonly used tubing materials for water isotopic studies – perfluoroalkoxy (PFA), fluorinated ethylene propylene (FEP), polytetrafluoroethylene (PTFE), high-density polyethylene (HDPE), and copper – at two different temperatures and an airflow rate of 0.635 L min−1 through approximately 100 ft (30.5 m) of 1/4 in. (6.4 mm) outer diameter (o.d.) tubing. All commonly used tubing materials performed similarly to each other in terms of attenuation times, reaching δ18O-location-adjusted δD and δ18O 95 % completion in less than 45 s, with slight variations based on temperature. PFA does appear to perform slightly better than the other materials, although memory metric differences are small. A tubing material commonly used in the early 2000s but reported to have memory effects on δD, Dekabon, was also tested at ambient temperature and changing humidities. The Dekabon isotopic equilibrium was not reached until nearly an hour after source transition, much later than H2O mixing ratios equilibrated. Bev-A-Line XX (used in some soil O2 and CO2 gas studies) was also tested at ambient temperature, but it did not approach isotopic equilibrium until after nearly 6 h of testing. Therefore, we cannot recommend the use of Bev-A-Line XX or Dekabon in water vapor isotope applications. Source transition from heavy to light or from light to heavy affected isotopic transition speed only in experiments where H2O ppmv was changing. While a shorter tubing lengths and smaller inner diameters shorten the delay of signal propagation through the tubing, they did not greatly change the attenuation curves under these conditions for the current commonly used tubing materials tested. However, in Dekabon, attenuation curves were greatly extended with increased tubing length. Our results show that the commonly used plastic tubing materials tested were not inferior to copper in terms of isotopic memory under these conditions, and they are easier to work with and are less expensive than copper.

Development of a forensic DNA research grade test material.

Advancements in forensic DNA typing technology and methods have increased sensitivity and, while beneficial, carry the weight of more challenging profile interpretation. In response, the forensic DNA community has often requested more complex reference materials to address commonly encountered measurement and interpretation issues such as complex DNA mixtures, DNA degradation, and PCR inhibition. The National Institute of Standards and Technology (NIST) released Research Grade Test Material 10235: Forensic DNA Typing Resource Samples to support the forensic DNA community. Components include three single source samples, two degraded samples, and three mixture samples. As part of the Research Grade Test Material (RGTM) process, automated methods for bottling, alternative sample tube types, and the addition of carrier RNA for stabilizing low-quantity samples were investigated. Both internal and external testing demonstrate the stability of the material over time at 4°C through qPCR testing. In the development of a data portal, users have been allowed to anonymously upload results and compare their data with NIST and others. This report describes the preparation and stability of this material.

Demonstration of potential DNA contamination introduced by laboratory consumables using Fluorescein

The development of increasingly efficient DNA extraction and profiling kits has increased the amount of allelic information obtained from trace DNA samples, but also inadvertently, increased the detection of DNA contamination. This study aimed to evaluate the potential of DNA transfer using fluorescein, fluorescent under an alternate light source, in the use of a range of forensically relevant DNA profiling consumables. An evaluation of two pre-lysis methods adopting three different sample tubes, some with deliberate seal damage, showed the PrepFiler™ Automated Forensic DNA Extraction Kit caused leakage and crusting when the rim of the PrepFiler™ LySep column was compromised, but no leakage was observed under the same conditions using the Investigator STAR Lyse&Prep kit. The AutoLys tube showed minimal leakage using the PrepFiler™ chemistry. A DNA extract tube with an external thread, similar to the AutoLys tube, showed no leakage after fridge or freezer storage. However, it highlighted that a centrifugal spin does not guarantee all the DNA will pool at the base of the tube. A comparison of adhesive plate sealing films to 8-well strip caps for sealing 96-well PCR plates showed the adhesive plate sealing films presented a lower risk of DNA transfer, largely due to the adhesion of dispersed liquid on the sticky surface of the film. Overall, this study highlighted a number of variables that may be considered in the development of more refined contamination minimisation protocols in respect to increased sensitivities of DNA profiling.

B-040 Direct Aspiration of Cryogenic Matrix Tubes on the Siemens Atellica System With Epic-Beaker Integration of External Barcode IDs for Analysis Small Volume Biobanked Specimens

Abstract Background Most core chemistry and immunochemistry clinical analyzers are designed for direct sampling of primary blood tubes with volumes ranging from 3.5-10 mL. This limits analysis of low volume pediatric specimens and biobanked aliquoted samples from clinical trials without manual technologist intervention. This work outlines a microsampling and data transmission scheme for analysis of 500 uL aliquots of biobanked serum collected for the Nutrition for Precision Health study powered by the NIH All of Us Research Program without relabeling secondary aliquot tubes after receipt. Methods Commercial plastic clip bearings sourced from IGUS (LCM-08-02) were used to stabilize the matrix tube in a standard 5 mL Sarstedt aliquot tube. 96-well format Matrix Tubes (ThermoFisher 3719-11) with pre-printed 10-digit side barcodes were programmed in the Atellica software as a custom false bottom tube with specifications height 75 mm, diameter 12 mm, bottom offset 41 mm and were loaded using a dedicated false bottom tray. Manifest files (.csv) containing the sample ID, clinical trials participant ID, collection date/time, age and sex were ingested into Epic via an Incoming Orders HL7 interface. Non-human patient records were generated using the participant ID as an external patient ID and patient name and calculating the Date of Birth based on the age in years given in the manifest file. Test orders were created based on assigned testing for the sample ID in the manifest. The sample ID corresponding the preprinted Matrix Tube barcode ID was inserted into the Epic container identifier field. Results Test orders were automatically transmitted to the Atellica Data Manger (ADM) allowing specimens to be run on the Atellica System (CH and IM modules) and results transmitted to Epic utilizing the established Data Innovations (DI) interface and processes used for patient samples without the need for Epic generated labels. Successful scanning and recognition of Matrix Tube barcodes was 100%. Complete aspiration of a Comprehensive Metabolic Panel, Lipid Panel, Iron, TIBC, hsCRP, insulin, and H, I, L indices occurred for 50% of samples tested without intervention. Approximately 250 uL of the 500 uL aliquot remained after aspiration of all tests (247.7 uL total volume) from the Matrix Tube. No probe crashes were observed despite the narrower 8 mm diameter of the Matrix Tubes versus the standard 12- or 13-mm diameter of primary blood collection tubes. Conclusions This Epic-DI-ADM data workflow and microsampling adaptation for Matrix Tube aspiration allows greater container type flexibility for the receipt and analysis of external samples delivered in non-primary blood tubes for clinical laboratories. Future work involving adjustment of the false bottom tube offset depth may reduce unusable sample volume allowing for a higher percentage of complete aspiration without intervention. This work is significant because it suggests existing clinical analyzers are easily adaptable to microsampling smaller volume tubes without significant effort.

A-122 Evaluation of the Performance of the BD MiniDraw™ Capillary Blood Collection System Under Various Transport Conditions

Abstract Background Specimen transport is a vital component of the preanalytical phase. External factors such as temperature fluctuations, vibrations, shock, and prolonged transport times may negatively impact specimen integrity and compromise test results. A study was performed to evaluate the effects of temperature extremes and simulated transportation on specimen quality in the BD MiniDraw™ Capillary Blood Collection System with BD MiniDraw™ H&H and SST™ Capillary Blood Collection Tubes (BD MiniDraw™ H&H, BD MiniDraw™ SST™). The BD MiniDraw™ Capillary Blood Collection System enables capillary blood collection by trained healthcare workers in non-traditional settings (i.e., retail pharmacies and clinics). Methods Sixteen adult participants were enrolled from the on-site Associate Sample Collection Program. Venous blood was collected via venipuncture into reference comparator and no-additive tubes. Venous blood from no-additive tubes was then pooled into a secondary container for aliquoting into BD MiniDraw™ SST™ and MiniDraw™ H&H tubes. Thirty tubes per tube type were separated into 6 groups with storage and simulated transport conditions conducted in accordance with ASTM D7386-16 and ASTM D4169-22, as applicable: Group 1a: No transport, 2-8°C storage uncontrolled humidity, 49-52-hour storage; Group 1b: Air/road transport, 2-8°C storage uncontrolled humidity, 49-52-hour storage; Group 2a: No transport, 20-24°C storage, uncontrolled humidity, 5-7-hour storage; Group 2b: Road transport, 20-24°C storage, uncontrolled humidity, 5-7-hour storage; Group 3a: Road transport, 38-40°C storage, 90% relative humidity, 5-7-hour storage; Group 3b: Air/road transport, 38-40°C storage, 90% relative humidity, 49-52-hour storage Following clotting, centrifugation, and storage for the BD MiniDraw™ SST™ tubes, testing was performed for selected chemistry analytes (Albumin, Alanine Aminotransferase (ALT), Alkaline Phosphatase (ALKP), Blood Urea Nitrogen, Chloride, Calcium, Cholesterol, Creatinine, High Density Lipoprotein (HDL), Low Density Lipoprotein (LDL), Sodium, Total Bilirubin (TBIL), Total Protein, Triglycerides) on the Siemens Atellica® Solution. Testing was performed for hemoglobin/hematocrit on the Sysmex XN-1000 in the BD MiniDraw™ H&H Tubes. Visual quality (barrier formation, hemolysis) and tube integrity (physical damage, blood leakage, cap removal, tube fit into adaptor) were also evaluated. Mean biases and confidence intervals within the specific analyte clinical acceptance limit for each group comparison was considered clinically equivalent. Results Clinical equivalence was demonstrated for all analytes for each group comparison except ALKP, ALT, HDL, LDL, TBIL and hematocrit for the extreme transport/storage conditions for Group 3b versus Group 3a. Complete barrier formation with acceptable visual hemolysis was noted in all specimens post transport. Tube integrity was maintained post transport as well. Conclusions Analytes were not impacted across a wide range of temperature and transport conditions. However, exposing samples to extreme high temperature, high humidity conditions may affect analyte performance. This reiterates the importance of monitoring conditions to ensure accurate test results. No impact to visual observations of sample quality and tube physical integrity was noted at different storage and transportation extremes.

B-207 Clinical Evaluation of ExiStation™ FA 96 RV for the detection of human respiratory viruses in human nasopharyngeal swab specimen

Abstract Background There has been growing demand for fully automated multiplex testing of respiratory viruses in molecular diagnostics following the COVID-19 pandemic. ExiStation™ FA 96 RV developed by BIONEER is a multiplex real-time RT-PCR assay. It is designed for the simultaneous detection and differentiation of SARS-CoV-2, Influenza A, Influenza B, and RSV RNA in human nasopharyngeal swab specimens from individuals showing signs and symptoms of respiratory viral infection. ExiStation™ FA 96 automates the entire process from RNA extraction to real-time RT-PCR for up to 96 samples at once within 100 minutes. Moreover, the system is equipped to decap sample tubes, eliminating the need for handling sample transport media. In this study, the clinical performance of the ExiStation™ FA 96 RV assay was evaluated using nasopharyngeal swab samples. Methods Nasopharyngeal swab samples, totaling 859 samples, are collected according to the procedures by the clinical institution in France and preserved in Virus transport media. Once anonymized and randomly allocated, the swab transport media tube, without capping procedure, are directly placed onto the ExiStation™ FA 96. Subsequently, both extraction and real-time RT-PCR are automatically processed. Clinical sensitivity, specificity, and agreement are analyzed by comparing results with confirmatory reagents SARS-CoV-2 RT-qPCR Reagent Kit, PKAmp™ Respiratory SARS-CoV-2 RT-PCR Panel Kit, and Bosphore SARS-CoV-2/Flu/RSV Panel Kit, as well as comparator reagent Allplex™ Respiratory Panel 1 and Allplex™ SARS-CoV-2 Assay. Results The comparison between confirmatory reagents and the ExiStation™ FA 96 RV resulted in a clinical sensitivity of at least 96.4% (164/164 for SARS-CoV-2, 64/65 for Influenza A, 74/75 for Influenza B, and 53/55 for RSV) and a clinical specificity of 100% (500/500 for all targets). When compared with the comparator reagent, the overall agreement and Cohen's kappa value were both determined to be above 99.1% and 0.989, respectively (664/664 for SARS-CoV-2, 564/565 for Influenza A, 573/575 for Influenza B, and 550/555 for RSV). Additionally, any discrepant samples identified in comparison with the comparator reagent were verified through sequencing analysis. Upon sequencing analysis on a total of 8 samples and aligning the confirmed results with the analysis content, no alterations observed in the analysis of clinical sensitivity and specificity. Conclusions The evaluation of the ExiStation™ FA 96 RV assay showed outstanding clinical performance in detecting four respiratory viruses, SARS-CoV-2, Influenza A, Influenza B, and RSV, with high sensitivity and specificity. This study supports that the ExiStation™ FA 96 RV assay is a reliable tool for simultaneous molecular diagnostics of various respiratory infections in clinical laboratories.

B-357 Siemens CA 72-4: an assay for measurement of Tumor Associated Glycoprotein (TAG) 72

Abstract Background The tumor associated glycoprotein (TAG) 72, also known as CA 72-4 is a mucin protein of high molecular weight. CA 72-4 is found on the surface of many cancer cells, including stomach (gastric), ovary, breast, colon and pancreatic cells. CA 72-4 is currently one of the primary markers for the diagnosis of gastric cancer and monitoring the course and efficacy of gastric cancer. In ovarian cancer, it has a higher sensitivity for mucin- type ovarian cancer, and its combination with CA125 for detection can significantly improve the clinical sensitivity. In colorectal cancer, CA 72-4 combined with CEA detection was also found to significantly improve the sensitivity of the initial diagnosis. An assay for CA 72-4 on Siemens Atellica IM (Siemens CA 72-4) is being developed and analytical performance is being presented. Methods The Siemens CA 72-4 assay (in development) is a one-step chemiluminescent microparticle immunoassay (CMIA) for the quantitative determination of CA 72-4 in human serum and plasma. The specimen is incubated with paramagnetic microparticles coated with the monoclonal antibody (mAb) B72.3 and acridinium-labeled mAb CC49 conjugate on the Siemens Atellica IM® System. After incubation and washing, Acid and Base reagents are added. The resulting relative light units (RLUs) are directly proportional to the amount of CA 72-4 in the sample allowing for the quantitative determination of CA 72-4 in the specimen. Results The following performance were determined using 3 unique lots of reagents and calibrators with the calibration range for the assay being 0.00 to 200.00 U/mL. The limit of blank (LoB), limit of detection (LoD), and limit of quantitation at 20% CV (LoQ) were determined and met the goal of ≤ 1.00 U/mL. Linearity according to CLSI EP06 Ed2 was demonstrated for a range of 1.00 through 200.00 U/mL. A Precision study of 2 controls and 5 panels spanning the range of the assay demonstrated a total %CV ≤ 6% for samples ≥ 5.00 U/mL and ≤ 0.25 SD for samples <5.00 U/mL. In the sample tube type study, a matrix comparison of 23 matched serum and plasma samples were evaluated. Passing-Bablock slopes of < 8% and r > 0.99 were observed when evaluating samples within the measurement range for plasma (LiHep, LiHep separator, K2EDTA and K3EDTA) tubes and serum separator tubes (SST) compared to the Red Top serum samples. Five endogenous substances were evaluated for interference in the CA 72-4 assay. The average percent difference between test and control samples for all endogenous interferents was < 10%. Percent difference in the presence of 28 potentially interfering drugs was ≤ 4.0%. Conclusions The Siemens CA 72-4 assay currently in development is a sensitive and precise assay for the quantitative determination of CA 72-4 in human serum and plasma.

B-228 Performance Comparison of three viral RNA pathogens in MK Buffered Solution for Diagnostic Based Detection using quantitative PCR vs droplet digital PCR

Abstract Background Droplet digital PCR (ddPCR) by partitioning a single reaction into many thousands of nano-droplets allows for the absolute quantitation of a given DNA/RNA target and represents the gold standard for the identification and quantification of pathogens in clinical and environmental samples. While nucleic acid based molecular diagnostics have become fundamental for the accurate and rapid quantification of specific pathogens, these instruments are still based upon quantitative PCR (QPCR) assays. The goal of this study was to assess the compatibility of a MK Buffered solution with three different RNA viruses combined into two standard clinical negative matrices and compare performance of ddPCR and QPCR for all three pathogens. Methods Serial dilutions of heat-inactivated Sars-CoV-2 (NR-52286), Influenza A (ATCC VR-1469) and RSV-A (ATCC strain VR-26) were combined into a single tube and added to two confirmed clinically negative matrices, Nasal and Sputum (Lee Biosolutions, Maryland Heights, MO). Serial dilutions of the combined pathogens were added to sample tubes containing 500 µL of MK buffered solution yielding final concentrations of each pathogen over 4 orders of magnitude in each clinical matrix. For each sample 300 µl was processed for RNA isolation using standard RNA isolation kits (Zymo Viral RNA Kit). For ddPCR samples were quantified for all three pathogens using a BioRad® Droplet Digital PCR System (QXD200). For QPCR, samples were quantified using Quantitative real-time PCR using Promega GoTaq® Probe 1- Step RT-qPCR System. The same TAQMan® probes specific for each pathogen were used for both systems. Results Yields of all three viral RNAs were significantly lower from sputum samples as compared to nasal samples. For all three pathogens the ddPCR assay was an order of magnitude more sensitive than observed with QPCR. QPCR and ddPCR values showed a significant linear relationship for all three pathogens (Sars-CoV-2, r2 = 0.984; RSV A, r2 = 0.814; Influenza A, r2 = 0.985). Conclusions QPCR and Droplet digital PCR accurately quantified concentrations of all three RNA pathogens in both clinically negative nasal and sputum matrices across four orders of magnitude, though detected yields were significantly lower in samples spiked into the sputum matrix. For all three pathogens there was a significant linear relationship with decreasing concentration. ddPCR was an order of magnitude more sensitive than QPCR for all three pathogens.

B-356 Siemens CYFRA 21-1: an assay for measurement of cytokeratin fragment 21-1

Abstract Background Cytokeratins are structural proteins forming the subunits of epithelial intermediary filaments. The soluble fragment of cytokeratin 19, cytokeratin fragment 21-1 (CYFRA 21-1), can be found in the blood of lung cancer patients, primarily those with non-small cell lung cancer (NSCLC). CYFRA 21-1 is the preferred biomarker in the management of NSCLC, particularly squamous cell and large cell carcinoma subtypes. An assay for CYFRA 21-1 on Siemens ADVIA Centaur® XPT and Siemens Atellica® IM (Siemens CYFRA) is being developed and analytical performance is being presented. Methods The Siemens CYFRA 21-1 assay (in development) is a one-step chemiluminescent microparticle immunoassay (CMIA) for the quantitative determination of CYFRA 21-1 in human serum and plasma. The specimen is incubated with paramagnetic microparticles coated with the monoclonal antibody (mAb) BM19.21 and acridinium-labeled mAb KS19.1 conjugate on the Siemens ADVIA Centaur XPT System and Siemens Atellica IM System. After incubation and washing, Acid and Base reagents are added. The resulting relative light units (RLUs) are directly proportional to the amount of CYFRA 21-1 in the sample allowing for the quantitative determination of CYFRA 21-1 in the specimen. Results The following performance were determined using 3 unique lots of reagents and calibrators with the calibration range for the assay being 0.00 to 100.00 ng/mL. For the ADVIA Centaur XPT and Atellica IM systems were shown to be equivalent within 4%. The limit of blank (LoB), limit of detection (LoD), and limit of quantitation at 20% CV (LoQ) were determined and met the goal of ≤ 1.00 ng/mL. Linearity according to CLSI EP06 Ed2 was demonstrated for a range of 0.66 through 100.00 ng/mL. A Precision study of 2 controls and 5 panels spanning the range of the assay demonstrated a total %CV ≤ 8% at all levels. In the sample tube type study, a matrix comparison of 61 matched serum and plasma samples were evaluated. Passing-Bablock slopes of < 5% and r > 0.99 were observed when evaluating samples within the measurement range for plasma (LiHep, LiHep separator, K2EDTA and K3EDTA) tubes and serum separator tubes (SST) compared to the Red Top serum samples. Six endogenous substances were evaluated for interference in the CYFRA 21-1 assay. The average percent difference between test and control samples for all endogenous interferents was < 10%. Percent difference in the presence of 20 potentially interfering drugs was ≤ 4.0%. Conclusions The Siemens CYFRA 21-1 assay currently in development is a sensitive and precise assay for the quantitative determination of CYFRA 21-1 in human serum and plasma.

A-060 Assessing Glucose Stability: A Comparison of Citrate/NaF, NaF, and Gel Separator Vacuum Tubes in Routine Medical Laboratory Practice

Abstract Background In cases where a glycolysis inhibitor is not utilized, blood cells in vitro can consume glucose. Although current guidelines suggest using citrate-buffered inhibitors or ice-water slurry to mitigate this, practical and financial constraints often hinder adherence to these recommendations. Our objective was to evaluate the difference in glucose concentration in our routine sample management by employing Citrate buffered/NaF, NaF, and Gel Barrier tubes. Methods In this study, we utilized Greiner FCmix citrate-buffered NaF and EDTA tubes (3 mL), Becton Dickinson NaF and EDTA tubes (4 mL), and Becton Dickinson SSTII Gel Barrier tubes (5 mL). Following venipuncture, the tubes were transferred to the laboratory and centrifuged within 30 minutes. Glucose levels of 46 patients' samples were measured using a Cobas 6000 system (c501) employing the Glucose Hexokinase method, which exhibits an intermediate coefficient of variation (CV) of approximately 1.5% and no significant bias, as confirmed by monitoring with Biorad EQAS. Results The mean plasma glucose difference between Citrate-buffered tubes and NaF tube samples was 2.8 mg/dL with a 95% confidence interval (CI) ranging from 2.2 to 3.4 mg/dL (Mean: 0.16 mmol/L; CI: 0.12 - 0.19 mmol/L). The difference in paired samples of NaF plasma and Gel Barrier tubes serum glucose was 3.2 mg/dL with a CI of 2.5 - 3.9 mg/dL (Mean: 0.18 mmol/L; CI: 0.14 - 0.22 mmol/L). Lastly, the mean difference in Citrate-buffered plasma and serum Gel barrier tubes for 46 paired samples was 6.0 mg/dL with a CI of 5.2 - 6.7 mg/dL (Mean: 0.33 mmol/L; CI: 0.29 - 0.37 mmol/L). Conclusions Following guidelines, Citrate-buffered/NaF tubes demonstrate efficacy as glucose stabilizers. When compared to NaF tubes alone within a 30-minute timeframe, the mean difference between the two types of tubes was 2.8 mg/dL. Although NaF tubes are no longer recommended for glucose testing in plasma, they still outperform gel barrier serum tubes by 3.2 mg/dL. Furthermore, compared to citrate/NaF Plasma, serum glucose levels were on average 6.0 mg/dL lower. The preanalytical sample stage poses a significant obstacle to its reliable use for diagnostic or monitoring purposes. Laboratories should consider transitioning to Citrate/NaF tubes, or employ mathematical corrections if utilizing gel separator tubes for glucose testing in their established preanalytical process.

Failure analysis in pyrolysis furnaces: Impact of carburization and thermal cycles on tube properties

The dominant failure mechanism in radiation coil tubes in pyrolysis furnaces is the detrimental interaction between carburization and reduced material ductility. This combination results in localized deformations, significant ovalization, and cracking in the tubes. At the same time, a second critical failure mechanism emerges during emergency furnace shutdowns, characterized by brittle fractures that can generate extensive longitudinal cracks in multiple tubes. This work presents a case study on the occurrence of failures that resulted in brittle fracture rupture of several tubes in a pyrolysis furnace that accumulated 43,720 h of operation. This highlights the practical importance of monitoring and managing the integrity of coil tubes. The tube samples were analyzed using optical and scanning electron microscopy, evaluation of carburization through magnetic permeability and NACE Test TM498, identification of carbides by X-ray diffraction (XRD), and dilatometry. Additionally, thermal stress analyses were performed using the finite element method, along with tensile tests at different temperatures. It was found that the tubes of the coil that operated at lower temperatures in their operational cycle did not rupture during the emergency shutdown, unlike other coils whose tubes experienced significant failures. An important contribution from this study is the demonstration of optimized operational cycle management and thermal control are essential in preserving protective oxide layers, minimizing coke formation and the effects of carburization and creep. This can reduce shutdown frequency and maintenance costs, improving the cost-effectiveness of the cracking process.

CCL2 may contribute to the damage of fallopian tube epithelium in women with tubal endometriosis

Research QuestionHow does tubal endometriosis (TEM) impact the morphology and ultrastructure of the fallopian tube epithelium? What are the underlying pathophysiological mechanisms of TEM? DesignEpithelial samples of the fallopian tubes from patients with (TEM group) and without (non-TEM group) TEM were collected. The morphological characteristics, ultrastructure, ciliated cell percentage, and ciliary beat frequency (CBF) were assessed via hematoxylin-eosin staining, electron microscopy, and high-speed video microscopy. mRNA microarray analysis was conducted to identify differentially expressed genes (DEGs) in the fallopian tube epithelium, which were further validated using qPCR, immunohistochemistry, and Western blotting. The effects of recombinant CCL2 protein on primary human fallopian tube epithelial cells (FTEC) were examined in vitro. ResultsPatients with TEM exhibited significant abnormalities in the morphology and ultrastructure of the fallopian tube epithelium with reduced percentage of ciliated cells and CBF, compared to the non-TEM patients. Among the 765 DEGs identified in the fallopian tube epithelium, 512 genes were upregulated and 253 genes were downregulated in the TEM group. qPCR, immunohistochemistry, and Western blotting validation confirmed a significant upregulation of CCL2 expression in the TEM group. In vitro experiments revealed that recombinant CCL2 protein significantly reduced ciliated cell differentiation, length of cilia, and CBF in FTEC, suggesting a role for CCL2 in the development of the TEM-related pathological phenotype. ConclusionsThese findings indicate that CCL2 may contribute to the pathological phenotype associated with TEM and could be a crucial signaling molecule in the damage of fallopian tube epithelium in patients with TEM.

Measurement of atmospheric amines and aminoamides by column adsorption/extraction and hydrophilic liquid chromatography-electrospray-tandem mass spectrometry.

Sampling and chromatography-mass spectrometry methods were investigated to measure atmospheric amines and aminoamides. Amines and their amide derivatives play significant roles in new particle formation (NPF) in the atmosphere, especially diamines and aminoamides have higher NPF potentials compared to monoamines. For amine sampling, silica gel tube collection and formic acid extraction gave good overall recoveries (>93 ± 8%) for mono-, di-, tri-, tetramines, and aminoamides. Two chromatography methods were subjected to analyze the extracted amines. One involved direct analysis using hydrophilic interaction liquid chromatography with carboxyl or diol group functioned separation column (carboxyl-HILIC or diol-HILIC), and the other utilized derivatization with 4-(N,N-dimethylaminosulfonyl)-7-fluoro-2,1,3-benzoxadiazole (DBD-F) and subsequent reversed-phase chromatography (HPLC). Separated amines were detected by electrospray ionization and tandem mass spectrometry in both cases. DBD-F-HPLC method provided good sensitivity for mono- and all polyamines (limit of detection (LOD) < 4.6nM, relative standard deviation (RSD) for 100nM < 9.2%). However, aminoamides could not be detected by DBD-F-HPLC. Carboxyl-HILIC provided good sensitivities for mono- and diamines and aminoamides (LOD < 1.6nM, RSD < 4.8%). Forest air measurement was performed and data obtained by carboxyl-HILIC and DBD-F-HPLC showed good agreement for 1,3-diaminopropane, 1,4-diaminobutane (putrescine) and 1,5-diaminopentane (cadaverine) (R2 = 0.9215-0.9739, n = 7-14). Carboxyl-HILIC method was the best for the amine analysis, and combination with silica gel tube sampling provides atmospheric monitoring available. The developed method can be used not only to study atmospheric chemistry of diamines and aminoamides but also to analyze flavor/odor of foods, flowers and wastes.

Effects of Wildfire Smoke on Volatile Organic Compound (VOC) and PM2.5 Composition in a United States Intermountain Western Valley and Estimation of Human Health Risk

With a warmer and drier climate, there has been an increase in wildfire events in the Northwest US, posing a potential health risk to downwind communities. The Lewis–Clark Valley (LCV), a small metropolitan area on the Washington/Idaho border in the United States Intermountain West region, was studied over the time period of 2017–2018. The main objective was to determine the community’s exposure to particulate matter (PM2.5) and volatile organic compounds (VOCs) during wildfire smoke events and to estimate the associated health risk. VOCs were analyzed previously in the LCV using sorbent tube sampling and thermal-desorption gas-chromatography mass-spectrometry (TD-GC-MS) during several local smoke events in the 2017–2018 fire seasons. PM2.5 measurements were obtained from nearby agency monitors. PM2.5 reached up to 200 µg/m3 in 2017 and over 100 µg/m3 in 2018 in the LCV, and has been observed to be increasing at a rate of 0.10 µg m−3/yr over the past two decades. Benzene, a carcinogen and air toxic, was measured with concentrations up to 11 µg/m3, over ten times the normal level in some instances, in the LCV. The health risk in the LCV from benzene was calculated at seven extra cancers per million for lifetime exposure and thirteen extra cancers per million considering all air toxics measured. The other cities monitored showed similar lifetime cancer risk, due to benzene of about 6–7 extra cancers per million. This work is important, as it measures ground-level exposures of VOCs and demonstrates decreases in PM2.5 air quality over time in the region.

Generating pooled quality control samples of volatile organic compounds

Untargeted analysis of volatile organic compounds (VOCs) from exhaled breath and culture headspace are influenced by several confounding factors not represented in reference standards. In this study, we propose a method of generating pooled quality control (QC) samples for untargeted VOC studies using a split-recollection workflow with thermal desorption tubes. Sample tubes were desorbed and split from each sample and recollected onto a single tube, generating a pooled QC sample. This QC sample was then repeatedly desorbed and recollected with a sequentially lower split ratio allowing injection of multiple QC samples. We found pooled QC samples to be representative of complex mixtures using principal component analysis and may be useful in future longitudinal, multi-centre, and validation studies to assess data quality and adjust for batch effects.

A case study on the impact of coppicing on small mammal diversity: First evidence from the high Agri Valley in the Basilicata Region, Italy

Forestry seems to have a significant impact on the diversity and composition of forest small mammal communities. Coppicing is one of the most widely adopted silvicultural systems in Europe and its effect on the entire small mammals community remains unclear. Thus, our study aimed to preliminary assess the stand-level effect of different coppice woodland elements on community richness, focusing on forest small mammal species. Furthermore, a second aim was to assess the effect of these woodland stand elements on different small mammal taxonomic groups to evaluate possible differential responses within the community. We identified three different coppice age classes, and selected a total of 15 coppice-with-standards forest stands. Additionally, we collected data about understory shrub structure and composition. Fur samples were collected using the hair tube sampling method. We found a significant effect of the coppice age on the number of species detected in communities. Furthermore, we found a differential response between glirid and murid families, with glirid being positively affected by a higher shrub abundance, which is instead severely reduced by coppicing.

Improving sample preparation by biochar-coated sampling tubes: proof-of-concept extraction of sex hormones from real waters

This work showcases a novel application of biochar for analytical sample treatment. The carbonaceous material, obtained by pyrolysis of orange peel waste (550°C) without any post-synthesis treatment or functionalization, was thoroughly characterized and easily immobilized on the inner wall of sampling tubes in order to perform a sort of “in-vial” solid-phase extraction (SPE). The as-obtained device was tested for the extraction of seven sexual steroids, as probe water pollutants, from tap, lake, river water and wastewater treatment plant effluent samples spiked with 0.2-5 µg L−1 of each compound. The sorption kinetics profiles showed quantitative uptake from the sample (25 mL) in 20 min contact time, followed by complete elution in pure ethanol (2 mL, 15 min), thanks to the proper balance between sorption affinity and ease of elution. Under the selected conditions, recovery was in the range 60-123 %, with good inter-day precision (RSD 10-18 %, n=3). As evidence of the excellent reproducibility, an overall RSD below 15 % was observed from inter-day inter-batch recovery tests on three individually prepared sampling tubes. The procedure, carried out on a roller mixer, allows 10 samples to be extracted simultaneously, improving the sample throughput. Moreover, reusability tests showed that the same device maintains its efficiency for 10 consecutive sorption/desorption cycles. The greenness assessment, carried out by two dedicated software, further supported the sustainability of this biochar-based sample preparation as an alternative SPE.

27 Effects of particle size and trace mineral source on vitamin stability in premix

Abstract Trace minerals and vitamins are necessary in diets for multiple physiological processes related to structural development, catalytic and regulatory systems in the animal. They become especially important when animals are challenged. These ingredients are commonly added to commercial diets through a concentrated mix, a premix. Unfortunately, vitamins are relatively unstable, and minerals are highly reactive, leading to reduction of the vitamins by the minerals. Feeding a mineral of a different particle size can influence this mechanic, as well as a different source, for example hydroxy trace minerals. The objective of this work was to evaluate vitamin A stability in presence of different sources of trace minerals as well as premix conditions. A microcrystalline cellulose matrix was created by dissolving vitamin A in an organic solvent (IPA). A fixed amount of vitamin MCC mix was added to a sample tube per analysis, containing common premix vitamin and mineral levels. Fixed amount of moisture and antioxidant (BHT) was added to evaluate these effects. Results on the effect of moisture indicated that 10% moisture addition had a large effect on vitamin A stability, reducing it by over 50% after 11 d. Vitamin A loss in premixes containing no moisture showed no difference between MCC mixes containing no mineral and ones containing hydroxy Cu, whereas there was a significant drop in vitamin A with CuSO4. Vitamin A stability was significantly affected by a lower particle size Cu source, with CuSO4 below 75 µm showed almost full degradation after 13 d. Hydroxy Cu showed relatively low degradation for its particle size. Overall, the results showed that moisture and particle size can have a huge impact on vitamin A stability, with BHT or antioxidants only partly being able to reduce this. Feeding a higher quality trace mineral source such as hydroxy trace minerals lead to a less reactive Cu, and hence, a higher vitamin A stability.

Solvent‐free crystallization for fractionation of virgin coconut oil: Effect of process conditions on kinetics and crystal properties

AbstractThis research was aimed to conduct solvent‐free crystallization and fractionation of virgin coconut oil (VCO). This study investigated the effect of surface contact area‐to‐volume ratio and mixing conditions on crystal characteristics. Commercially available VCO samples, extracted by different methods, were analyzed for fatty acid and triacylglycerol composition using GC–MS and UPLC–ELSD. VCO extracted through enzyme and chilling–thawing cycles exhibited higher medium chain fatty acids and medium chain triacylglycerols, which was selected for crystallization studies. Isothermal crystallization of VCO at 21°C assessed the effect of scale of crystallizer tube on cooling rate using Newton's law of cooling equation. The smaller tube (20 mm diameter) with a higher surface contact area‐to‐volume ratio produced 4.1‐fold faster cooling and a higher crystallization yield (44%) compared to the 80 mm diameter tube when no agitation was applied. Whereas mixing with a magnetic stirrer produced a large number of nuclei, generating intense crystallization heat, and rendering the crystals inseparable from liquid fraction. The process of VCO crystallization was also studied for kinetics and crystal growth mechanism using the Avrami model employing angled rotation of sample tube during non‐isothermal crystallization. Angled rotary mixing produced stable crystals and significantly increased the crystallization rate (t1/2 = 26.86 min) with polyhedral or spherulitic crystal growth (n = 3.25) compared to non‐mixing condition (t1/2 = 161.87 min) with restricted linear growth (n = 1.64). DSC analysis of VCO and its fractions obtained after crystallization using the 20 mm tube with angled rotary mixing indicated shifts in their thermogram peaks, suggesting differences in triacylglycerol compositions.Practical applicationsVirgin coconut oil (VCO) is well recognized for its health‐beneficial properties, yet its application is limited due to its complex triglyceride composition. The solvent‐free crystallization method facilitates VCO fractionation into phases with different melting points without hazardous solvents. The efficacy of this process is highly dependent on temperature, cooling rate, and agitation, which affects crystal morphology and growth kinetics. This study addresses these challenges in the scarcely investigated area of VCO crystallization. The resulting fractions can be customized for specific applications based on their altered functional properties in producing baking and confectionery coatings, salad dressings, and so on. The olein fraction obtained in this study can be utilized in nutraceutical formulations targeting digestive and cognitive health, as it is reported to be rich in medium chain triglycerides. This research optimizes process conditions, suggesting an efficient method for dry‐fractionating VCO, which is significantly associated with the food and pharmaceutical industries.