Hematocrit-Independent Sampling Enables White Blood Cell Counts from Patterned Dried Blood Spot Cards.

The accurate and efficient measurement of white blood cell (WBC) counts is vital for monitoring general patient health and can aid in the diagnosis of a range of possible infections or diseases. Even with their importance universally acknowledged, access to WBC counts is largely limited to those with access to phlebotomists and centralized clinical laboratories, which house the instruments that perform the tests. As a result, large populations of people (e.g., those that are home-bound or live in remote locations) lack facile access to testing. Dried blood spot (DBS) cards are often used to bridge these gaps in access to testing by offering the ability to collect blood at home for ambient shipping to laboratories. However, it is well understood that these cards, which are prepared from cellulose cardstocks without further modification, suffer from variabilities in accuracy and precision due to uncontrolled sample spreading and hematocrit effects, which have hindered their use to determine WBC counts. In this paper, we present a method to obtain an accurate WBC count using a patterned dried blood spot (pDBS) card, which comprises collection zones that meter volumes of dried blood. Using an input volume of 75 µL of whole blood, we demonstrate that, unlike the gold standard DBS card (Whatman 903), our pDBS design allows for the collection of replicate zones containing a reproducible, average volume of dried blood (12.1 µL, 7.8% CV) over the range of hematocrits from 25 to 55%. We then used qPCR to quantify the 18S rRNA gene to determine WBC counts from the volumes of blood that are metered in pDBS zones. We observe that WBC counts generated from our method are comparable to those measured with a HemoCue point-of-care WBC analyzer. Our approach to using pDBS cards as a blood collection device has the potential to support at-home sampling and other patient populations that need WBC counts but lack access to clinical facilities.

Satisfaction with home blood sampling methods and expectations for future point-of-care testing in phenylketonuria: Perspectives from patients and professionals.

INTRODUCTION: Phenylketonuria (PKU) requires regular phenylalanine monitoring to ensure optimal outcome. However, home sampling methods used for monitoring suffer high pre-analytical variability, inter-laboratory variability and turn-around-times, highlighting the need for alternative methods of home sampling or monitoring. METHODS: A survey was distributed through email and social media to (parents of) PKU patients and professionals working in inherited metabolic diseases in Denmark, The Netherlands, and United Kingdom regarding satisfaction with current home sampling methods and expectations for future point-of-care testing (POCT). RESULTS: 210 parents, 156 patients and 95 professionals completed the survey. Countries, and parents and patients were analysed together, in absence of significant group differences for most questions. Important results are: 1) Many patients take less home samples than advised. 2) The majority of (parents of) PKU patients are (somewhat) dissatisfied with their home sampling method, especially with turn-around-times (3-5 days). 3) 37% of professionals are dissatisfied with their home sampling method and 45% with the turn-around-times. 4) All responders are positive towards developments for POCT: 97% (n = 332) of (parents of) patients is willing to use a POC-device and 76% (n = 61) of professionals would recommend their patients to use a POC-device. 5) Concerns from all participants for future POC-devices are costs/reimbursements and accuracy, and to professionals specifically, accessibility to results, over-testing, patient anxiety, and patients adjusting their diet without consultation. CONCLUSION: The PKU community is (somewhat) dissatisfied with current home sampling methods, highlighting the need for alternatives of Phe monitoring. POCT might be such an alternative and the community is eager for its arrival.

Effect of Minimization of Early Blood Sampling Losses Among Extremely Premature Neonates: A Randomized Clinical Trial.

OBJECTIVE: To evaluate the effect of blood sampling stewardship on transfusion requirements among infants born extremely preterm. STUDY DESIGN: In this single-center, randomized controlled trial (RCT), infants born at <28 weeks of gestation and birth weight of <1000 g were randomized at 24 hours of age to two different blood sampling approaches: restricted sampling (RS) vs conventional sampling (CS). The stewardship intervention in the RS group included targeted reduction in blood sampling volume and frequency and point of care testing methods in the first 6 weeks after birth. Both groups received early recombinant erythropoietin from day three of age. Primary outcome was the rate of early red blood cell (RBC) transfusions in the first six postnatal weeks. RESULTS: A total of 102 infants (mean gestational age: 26 weeks; birth weight: 756 g) were enrolled. Fidelity to the sampling protocol was achieved in 95% of the infants. Sampling losses in the first 6 weeks were significantly lower in the RS group (16.8 ml/kg vs 23.6 ml/kg, P < .001). The RS group had a significantly lower rate of early postnatal RBC transfusions (41% vs 73%, RR: 0.56 [0.39-0.81], P = .001). The hazard of needing a transfusion during neonatal intensive care unit (NICU) stay was reduced by 55% by RS. Mortality and neonatal morbidities were similar between the two groups. CONCLUSION: Minimization of blood sampling losses by approximately one-third in the first 6 weeks after birth leads to substantial reduction in the early red blood cell transfusion rate in infants born extremely preterm and weighing <1000 g at birth. TRIAL REGISTRATION: http://www.ctri.nic.in (CTRI/2020/01/022  964).

Microsampling Techniques Suitable for Therapeutic Drug Monitoring of Antipsychotics.

BACKGROUND: Therapeutic drug monitoring (TDM) of antipsychotics for dose titration or detection of noncompliance is not uncommon in daily practice. Normally, TDM implies measuring a drug concentration in venous blood samples. This technique is invasive and requires trained assistants and patients normally need to go to an outpatient clinic. Over the past decades, sensitivity of analytical equipment has improved leading to a growing interest in microsampling techniques. These techniques are minimally invasive, require a small volume (<100 µL), usually result in stable samples, and can be collected by the patient or a caregiver at home. Before a microsampling technique can be used in daily routine, proper method development and a clinical validation study should be performed. METHOD: For this review, the databases of PubMed and Embase were systematically searched. Currently available microsampling techniques for antipsychotics in blood, serum, or plasma are summarized. Subsequently, it has also been assessed whether these techniques are sufficiently validated for TDM monitoring in daily practice. RESULTS: Several microsampling techniques are available today, for example, dried blood spot sampling, dried plasma extraction cards, and volumetric absorptive microsampling. Eighteen studies were identified in which a microsampling technique for 1 or a few antipsychotics was chemically analytically and clinically validated. However, the majority of these studies have relevant shortcomings that mean its usefulness for different antipsychotics is not yet well established. CONCLUSIONS: Microsampling for TDM can be recommended for patients using clozapine. For TDM of other antipsychotics, it is a very promising development.

Novel method for determining when a field-collected donor unit is sufficiently full.

BACKGROUND: Whole blood (WB) collections can occur downrange for immediate administration. An important aspect of these collections is determining when the unit is sufficiently full. This project tested a novel method for determining when a field collection is complete. METHODS: The amount of empty space at the top of WB units, destined to become LTOWB or separated into components, that were collected at blood centers or hospitals was measured by holding a WB unit off the ground and placing the top of a piece of string where the donor tubing entered the bag. The string was marked where it intersected the top of the column of blood in the bag and measured from the top. The WB units were also weighed. RESULTS: A total of 15 different bags, two of which were measured in two different filling volumes, from 15 hospitals or blood centers were measured and weighed. The most commonly used blood bag, Terumo Imuflex SP, had a median string length of 9 mm (range: 2-24 mm) and weighed a median of 565.1 g (range: 524.8-636.7 g). CONCLUSION: Pieces of string can be precut to the appropriate length depending on the type of bag before a mission where field WB collections might be required and a mark placed on the bag before the collection commences to indicate when the unit is full.

Diagnostic sample transport via pneumatic tube systems: data logger and their algorithms are sensitive to transport effects.

OBJECTIVES: Many hospitals use pneumatic tube systems (PTS) for transport of diagnostic samples. Continuous monitoring of PTS and evaluation prior to clinical use is recommended. Data loggers with specifically developed algorithms have been suggested as an additional tool in PTS evaluation. We compared two different data loggers. METHODS: Transport types - courier, conventional (cPTS) and innovative PTS (iPTS) - were monitored using two data loggers (MSR145® logger, CiK Solutions GmbH, Karlsruhe, Germany, and a prototype developed at the University Medicine Greifswald). Data loggers differ in algorithm, recording frequencies and limit of acceleration detection. Samples from apparently healthy volunteers were split among the transport types and results for 37 laboratory measurands were compared. RESULTS: For each logger specific arbitrary units were calculated. Area-under-the-curve (AUC)-values (MSR145®) were lowest for courier and highest for iPTS and increased with increasing recording frequencies. Stress (St)-values (prototype logger) were obtained in kmsu (1,000*mechanical stress unit) and were highest for iPTS as well. Statistical differences between laboratory measurement results of transport types were observed for three measurands sensitive for hemolysis. CONCLUSIONS: The statistical, but not clinical, differences in the results for hemolysis sensitive measurands may be regarded as an early sign of preanalytical impairment. Both data loggers record this important interval of beginning mechanical stress with a high resolution indicating their potential to facilitate early detection of preanalytical impairment. Further studies should identify suitable recording frequencies. Currently, evaluation and monitoring of diagnostic sample transport should not only rely on data loggers but also include diagnostic samples.

Recommendations for blood sampling in emergency departments from the European Society for Emergency Medicine (EUSEM), European Society for Emergency Nursing (EuSEN), and European Federation of Clinical Chemistry and Laboratory Medicine (EFLM) Working Group for the Preanalytical Phase. Executive summary.

AIM: Blood Sampling Guidelines have been developed to target European emergency medicine-related professionals involved in the blood sampling process (e.g. physicians, nurses, phlebotomists working in the ED), as well as laboratory physicians and other related professionals. The guidelines population focus on adult patients. The development of these blood sampling guidelines for the ED setting is based on the collaboration of three European scientific societies that have a role to play in the preanalytical phase process: EuSEN, EFLM, and EUSEM. The elaboration of the questions was done using the PICO procedure, literature search and appraisal was based on the GRADE methodology. The final recommendations were reviewed by an international multidisciplinary external review group. RESULTS: The document includes the elaborated recommendations for the selected sixteen questions. Three in pre-sampling, eight regarding sampling, three post-sampling, and two focus on quality assurance. In general, the quality of the evidence is very low, and the strength of the recommendation in all the questions has been rated as weak. The working group in four questions elaborate the recommendations, based mainly on group experience, rating as good practice. CONCLUSIONS: The multidisciplinary working group was considered one of the major contributors to this guideline. The lack of quality information highlights the need for research in this area of the patient care process. The peculiarities of the emergency medical areas need specific considerations to minimise the possibility of errors in the preanalytical phase.

The stability of 65 biochemistry analytes in plasma, serum, and whole blood.

OBJECTIVES: The pre-analytical stability of various biochemical analytes requires careful consideration, as it can lead to the release of erroneous laboratory results. There is currently significant variability in the literature regarding the pre-analytical stability of various analytes. The aim of this study was to determine the pre-analytical stability of 65 analytes in whole blood, serum and plasma using a standardized approach. METHODS: Blood samples were collected from 30 healthy volunteers (10 volunteers per analyte) into five vacutainers; either SST, Li-heparin, K2-EDTA, or Na-fluoride/K-oxalate. Several conditions were tested, including delayed centrifugation with storage of whole blood at room temperature (RT) for 8 h, delayed centrifugation with storage of whole blood at RT or 4 °C for 24 h, and immediate centrifugation with storage of plasma or serum at RT for 24 h. Percent deviation (% PD) from baseline was calculated for each analyte and compared to the maximum permissible instability (MPI) derived from intra- and inter-individual biological variation. RESULTS: The majority of the analytes evaluated remained stable across all vacutainer types, temperatures, and timepoints tested. Glucose, potassium, and aspartate aminotransferase, among others, were significantly impacted by delayed centrifugation, having been found to be unstable in whole blood specimens stored at room temperature for 8 h. CONCLUSIONS: The data presented provides insight into the pre-analytical variables that impact the stability of routine biochemical analytes. This study may help to reduce the frequency of erroneous laboratory results released due to exceeded stability and reduce unnecessary repeat phlebotomy for analytes that remain stable despite delayed processing.

End-user verification results of two serum separator tubes for clinical chemistry analytes according to CLSI GP34-A and CLSI GP41-A6.

Tube manufacturers use different composition of gels and blood clot activator formulations in serum tube production. Our aim was to investigate the within-tube (repeatability) and between-tube variation, concordance between comparison results of BD and VacuSEL tubes. Blood samples were collected from control subjects (n = 20) and patients (n = 30) in accordance with the CLSI GP41-A6 and CLSI GP34-A guidelines. Twenty-three clinical chemistry parameters were analysed via Roche Cobas C702 Chemistry Analyzer on T0 (0 hour) and T24 (24 hour). Mean differences % were compared with Wilcoxon matched pair test. Clinical significance was evaluated based on desirable bias according to total allowable error (TEa). VacuSEL tubes demonstrated acceptable performance for the results of 20 parameters with regards to desirable bias % limits. Lactate dehydrogenase (LD) [mean difference % (%95 confidence intervals (CI) values of BD and VacuSEL tubes at T0 [6.41% (4.80-8.01%)]; sodium (Na) and total protein (TP) at T24 [-0.27% (-0.46 to -0.07%) and -1.39% (-1.87 to -0.91), respectively] were over the desirable bias limits (LD: 4.3%, Na: 0.23% and TP: 1.36%, respectively) but not exceeding total biological variation CV % [Na: 0.5 (0.0-1.0) % and TP: 2.6 (2.3-2.7) %). %95 confidence intervals (CI) of T0 LD values overlap with within-subject biological variation % (CI) limits (LD: 5.2 (4.9-5.4) %). The differences between two tubes were not medically significant and necessarily conclusive. VacuSEL serum tubes presented comparable performance with BD serum tubes.

Five days serum glucose stability at room-temperature in centrifuged fast-clotting serum tubes and the comparability with glucose in heparin-plasma and plasma containing citrate-stabilizer.

Glucose measurement plays a central role in the diagnosis of gestational diabetes mellitus (GDM). Because of earlier reports of overestimation of glucose in the widely used tubes containing granulated glycolysis inhibitor, the study assessed the performance of fast-clotting serum tubes as an alternative sample for the measurement of glucose. Glucose concentration in fast-clotting serum was compared to lithium-heparin plasma placed in an ice-water slurry after sample collection and glucose stability at room-temperature was studied. Blood samples from 30 volunteers were drawn in four different types of tubes (serum separator tubes, fast-clotting serum tubes, lithium-heparin tubes and sodium fluoride, EDTA and a citrate buffer (NaF-EDTA-citrate) tubes, all from Greiner Bio-One). Lithium-heparin tubes were placed in an ice-water slurry until centrifugation in accordance with international recommendations and centrifuged within 10 min. After centrifugation, glucose was measured in all tubes (timepoint T0) and after 24, 48, 72, 96 and 120 h of storage at 20-22 °C. NaF-EDTA-citrate plasma showed significant overestimation of glucose concentration by 4.7% compared to lithium-heparin plasma; fast-clotting serum showed glucose concentrations clinically equivalent to lithium-heparin plasma. In fast-clotting serum tubes, mean bias between glucose concentration after 24, 48, 72, 96 and 120 h and T0 was less than 2.4%. All individual differences compared to T0 were less than 6.5%. The results fulfill the acceptance criteria for sample stability based on biological variation. Fast-clotting serum tubes can be an alternative for the measurement of glucose in diagnosis and management of GDM and diabetes mellitus, especially when prolonged transportation is necessary.

Adrenal venous sampling criteria for chemiluminescent enzyme immunoassay as a preferable alternative to radioimmunoassay in primary aldosteronism.

Plasma aldosterone concentration (PAC) was routinely measured using radioimmunoassay (RIA); however, the RIA kit was discontinued in March 2021 in Japan. This study examined PAC conversion in adrenal venous sampling (AVS) and AVS criteria when measured using chemiluminescent enzyme immunoassay (CLEIA). PAC of 415 adrenal venous blood samples from AVS (including segmental AVS) of 63 patients with primary aldosteronism was measured using RIA (Spac-S aldosterone kit; Fujirebio Inc.) and CLEIA (Lumipulse Presto Aldosterone; Fujirebio Inc.). PAC of 70 AVS samples was also measured using liquid chromatography-mass spectrometry (LC-MS/MS, ASKA Pharma Medical Co., Ltd.). PAC conversion formulas were determined for each AVS sample assay. PAC measured using CLEIA was significantly correlated with that measured using RIA (correlation coefficient = 0.971). The PAC conversion formula was PAC (CLEIA) = PAC (RIA) × 0.772 - 1,199 pg/mL. The PAC of 14,000 pg/mL in RIA was equivalent to 9,613 pg/mL in CLEIA. PAC measured using CLEIA was also correlated with that measured using LC-MS/MS, and the PAC conversion formula was PAC (CLEIA, pg/mL) = 0.97 × PAC (LC-MS/MS, pg/mL) + 211. The inter-assay coefficient of variability (CV) was 1.1-1.3% and intra-assay CV was 1.0-1.7%, measured using CLEIA. The PAC conversion formula for AVS samples was obtained using CLEIA and RIA, and the conversion formula was different from that for peripheral blood. PAC values measured by CLEIA showed preferable accuracy and high concordance with those measured by LC-MS/MS, even in AVS samples. The study outcomes are useful for interpreting AVS results using non-RIA measurement methods.

The effect of the simulated intrauterine sound on behavioral and physiological indices of pain during capillary blood sampling for screening preterm infants: a randomized clinical trial study.

INTRODUCTION: Due to medical procedures, preterm infants are at high risk for side effects of pain. In this regard, heel lancing for capillary blood sampling is a common painful procedure. The present study was conducted to assess the effectiveness of a simulated intrauterine sound on behavioral and physiological indices of pain due to heel-prick blood sampling in preterm infants. METHODS: A double­blind randomized clinical trial (RCT) was conducted. The data were collected from September 23 to December 22, 2019. We measured the effect of a simulated intrauterine sound on changes in the behavioral and physiological parameters of pain (heart rate, SPO2) caused by heel lance that was measured 5 min before the intervention, during the sampling, and 5 min after the procedure. We measured behavioral pain by video recording the infants' faces and then the scoring neonatal infant pain scale (NIPS). Heart rate and SPO2 were measured using a pulse oximeter device. The data were analyzed using analysis of variance (ANOVA) and independent t­test in SPSS software version 20.0. RESULTS: Eighty infants were randomized (40 in each group). Mean scores NIPS during and after intervention were in the intervention group (3.55 ± 0.84, 95% CI: 3.30-3.80(, and (1.15 ± 0.84, 95%: 0.95-1.35) and in the control group (5.57 ± 0.95, 95% CI:5.30-5.85) and (3.00 ± 0.98) respectively. There were significant differences in scores of NIPS between the two study groups during (p < 0.001) and five min after heel lancing (p < 0.001). Mean scores of heart rate in the three phases of before, during, and five min after the intervention were respectively in the intervention group (127.57 ± 4.45, 95% CI:126.27-128.99), (131.07 ± 6.54, 95% CI:129.20-133.22), (128.45 ± 5.15, 95% CI:127.02-130.07) and in the control group (128.67 ± 4.57, 95% CI:127.32-130.07), (136.07 ± 7.24, 95% CI:133.90-138.37), and (132.42 ± 6.47, 95% CI:130.37-134.49). There were significant differences in heart rate between the intervention and the control group during (p = 0.002) and five min after the heel lance (p = 0.003). Mean scores of SPO2 in the three phases of baseline, during, and five min after the intervention were respectively in the intervention group (96.72 ± 0.93, 95% CI:96.42-97.00), (91.47 ± 1.46, 95% CI:91.05-91.92), (94.17 ± 1.03, 95% CI:93.22-94.00) and in the control group (96.6 ± 0.84, 95% CI:96.35-96.85), (91.5 ± 1.24, 95% CI:91.12-91.87), and (93.60 ± 1.27, 95% CI:93.85-94.50). CONCLUSION: This study showed that the simulated intrauterine sound reduces the behavioral pain and heart rate in the intervention group during and after heel lance. These results suggest using the method during the painful heel lancing to reduce pain parameters in preterm infants.

Impact of microsampling on toxicological evaluation in rodent safety studies.

Recently, animal welfare has been attracting worldwide attention, and implementation of 3Rs (replacement, reduction, and refinement) is prioritized in every way possible in the drug development. Microsampling, in which small amounts of blood are collected, is attracting attention in this context. ICH S3A Q&A focused on microsampling was published in November 2017 to help accelerate the application of microsampling for toxicokinetic assessment. The increased sensitivity of drug measurement apparatuses such as mass spectrometers has made it possible to measure drug concentrations with small amounts of blood samples. In this review, we summarized the reports on toxicological influence of microsampling in rodents (rats and mice) with or without drug administration or recovery period after blood collection and influences that may arise from differences in the blood sampling site or blood sampling volume. We also summarized some perspectives on further implementation of microsampling in toxicology studies. The use of microsampling in regulatory toxicology studies has gradually increased, although at a lower rate than in discovery studies. Since more animals are used in GLP toxicology studies than in discovery studies, the effect of reducing the number of animals by microsampling is expected to be greater in the toxicology studies. This report aims to promote the application of microsampling to nonclinical studies, as it is beneficial for improving animal welfare and can contribute to the 3Rs.

Short communication: Storage time and temperature affect plasma osmolality values in field-collected blood samples.

As climate change alters the hydric regime of many habitats, understanding the hydric physiology of animals becomes increasingly important. Plasma osmolality is a popular metric to assess an organism's hydration, but samples often need to be stored before being analyzed, under varying conditions and for different lengths of time. Previous studies on plasma storage conditions, and how they impact sample integrity, are minimal and have focused more on clinical applications than field studies. We studied the stability of osmolality values from wild rattlesnake plasma samples stored in commonly used plastic snap-cap tubes under different time (0, 2, 3, 7, 29 days) and temperature (refrigerated at 2 °C and frozen at -18 °C) treatments. We hypothesized that frozen samples would remain more stable (e.g., retain osmolality values more similar to baseline values) than refrigerated samples because freezing the plasma would reduce evaporation. We found that osmolality of samples increased over time at both temperatures, becoming significantly higher than baseline after 7 days. Contrary to our prediction, osmolality increased more in frozen samples than in refrigerated samples. We discuss possible reasons for our results, along with their implications. To obtain the most accurate plasma osmolality values, we recommend refrigerating plasma samples for as short a time as possible, 3 days or fewer, before analyzing them on an osmometer.

Comparison of laser and traditional lancing devices for capillary blood sampling in patients with diabetes mellitus and high bleeding risk.

PURPOSE: Despite the importance of self-monitoring blood glucose (SMBG) for management of diabetes mellitus (DM), frequent blood sampling is discouraged by bleeding risk due to dual-antiplatelet agent therapy (DAPT) or thrombocytopenia. METHODS: We compared the bleeding time (BT) of sampling by using a laser-lancing-device (LMT-1000) and a conventional lancet in patients with DM and thrombocytopenia or patients undergoing DAPT. BT was measured using the Duke method, and pain and satisfaction scores were assessed using numeric rating scale (NRS) and visual analog scale (VAS). The consistency in the values of glucose and glycated-hemoglobin (HbA1c) sampled using the LMT-1000 or lancet were compared. RESULTS: The BT of sampling with the LMT-1000 was shorter than that with the lancet in patients with thrombocytopenia (60s vs. 85s, P = 0.024). The NRS was lower and the VAS was higher in laser-applied-sampling than lancet-applied sampling in the DAPT-user group (NRS: 1 vs. 2, P = 0.010; VAS: 7 vs. 6, P = 0.003), whereas the group with thrombocytopenia only showed improvement in the VAS score (8 vs. 7, P = 0.049). Glucose and HbA1c sampled by the LMT-1000 and lancet were significantly correlated in both the DAPT-user and the thrombocytopenia groups. CONCLUSION: The LMT-1000 can promote SMBG by shortening BT in subject with thrombocytopenia and by increasing satisfaction score, as well as by showing reliable glucose and HbA1c value.

Establishing a plan to improve pediatric patient comfort during PIV insertions and blood specimen collection: a quality improvement effort.

HIGHLIGHTS: Patient comfort during peripheral intravenous (PIV) insertion and specimen collection was increased. The authors extended the contingency plan implemented for PICC insertion to include PIV insertion and specimen collection. The authors met their goals by using quality improvement methodology. Prioritizing patient comfort often requires institutional culture change. BACKGROUND: Needle procedures can cause pain and distress, especially in pediatric patients.1 Retrospective data collected at a freestanding pediatric facility revealed that approximately 30% of pediatric patients were not demonstrating sufficient levels of comfort during peripheral intravenous (PIV) catheter insertion and specimen collection (lab draws) even after successful implementation of comfort measures by the vascular access team (VAT) in an adjacent procedure (eg peripherally inserted central catheter placement). The current quality improvement project was implemented to support adaptation and expansion of previous lessons learned to PIVs and lab draws specifically. DESIGN AND METHODS: The VAT used the Pediatric Sedation State Scale,2 a standardized assessment tool integrated into the electronic medical record, to assess procedural comfort during PIVs and lab draws from February 2021 through April 2023. A total of 24 134 patients aged 0 to 18 years were included in the data collection. Interventions were delivered concurrently and included (1) reeducation/ongoing support for implementation of the Comfort Promise3 measures, (2) the creation and implementation of advanced comfort options, and (3) culture change. AIMS AND OBJECTIVES: The goal of the interventions was to improve the percentage of pediatric patients achieving adequate levels of comfort beginning at 68% in year 1 to 90% in year 2. RESULTS: From February 2021 to April 2023, the VAT team was able to improve procedural comfort scores from 68% to 90% of pediatric patients with adequate comfort for lab draws and/or PIV insertions. CONCLUSIONS: While standard comfort measures are a good first step in pain management during needle procedures, they are not sufficient for every pediatric patient. Nitrous, sedation, and the use of anxiolytics and analgesics can play an important role in reducing pain and anxiety during needle procedures and should be considered for patients not achieving adequate levels of comfort with standard comfort measures.

Semiautomated Electrochemical Melting Curve Analysis Device for the Detection of an Osteoporosis Associated Single Nucleotide Polymorphism in Blood.

The detection of single nucleotide polymorphisms (SNPs) is of increasing importance in many areas including clinical diagnostics, patient stratification for pharmacogenomics, and advanced forensic analysis. In the work reported, we apply a semiautomated system for solid-phase electrochemical melting curve analysis (éMCA) for the identification of the allele present at a specific SNP site associated with an increased risk of bone fracture and predisposition to osteoporosis. Asymmetric isothermal recombinase polymerase amplification using ferrocene labeled forward primers was employed to generate single stranded redox labeled amplicons. In a first approach to demonstrate the proof of concept of combining asymmetric RPA with solid-phase éMCA, a simplified system housing a multielectrode array within a polymeric microsystem, sandwiched between two aluminum plates of a heater device, was used. Sample manipulation through the microfluidic channel was controlled by a syringe pump, and an external Ag/AgCl reference electrode was employed. Individual electrodes of the array were functionalized with four different oligonucleotide probes, each probe equivalent in design with the exception of the middle nucleotide. The isothermally generated amplicons were allowed to hybridize to the surface-tethered probes and subsequently subjected to a controlled temperature ramp, and the melting of the duplex was monitored electrochemically. A clear difference between the fully complementary and a single mismatch was observed. Having demonstrated the proof-of-concept, a device for automated éMCA with increased flexibility to house diverse electrode arrays with internal quasi-gold reference electrodes, higher resolution, and broader melting temperature range was developed and exploited for the detection of SNP hetero/homozygosity. Using the optimized conditions, the system was applied to the identification of the allele present at an osteoporosis associated SNP site, rs2741856, in 10 real fingerprick/venous blood samples, with results validated using Sanger sequencing.

Assessment of a 60-Biomarker Health Surveillance Panel (HSP) on Whole Blood from Remote Sampling Devices by Targeted LC/MRM-MS and Discovery DIA-MS Analysis.

Telehealth, accessing healthcare and wellness remotely, should be a cost-effective and efficient way for individuals to receive care. The convenience of having a reliable remote collection device for blood tests will facilitate access to precision medicine and healthcare. Herein, we tested a 60-biomarker health surveillance panel (HSP), containing 35 FDA/LDT assays and covering at least 14 pathological states, on 8 healthy individuals' ability to collect their own capillary blood from a lancet finger prick and directly compared it to the traditional phlebotomist venous blood and plasma collection methods. All samples were spiked with 114 stable-isotope-labeled (SIL) HSP peptides and quantitatively analyzed by liquid chromatography-multiple reaction monitoring-mass spectrometry (LC/MRM-MS) scheduled method targeting 466 transitions from 114 HSP peptides and by a discovery data-independent acquisition mass spectrometry (DIA-MS) method. The average peak area ratio (PAR) of the HSP quantifier peptide transitions from all 8 volunteers' capillary blood (n = 48), venous blood (n = 48), and matched plasma (n = 24) was <20% coefficients of variation (CV). Heat map analysis of all 8 volunteers demonstrated that each individual had a unique biosignature. Biological replicates from capillary blood and venous blood clustered within each volunteer in k-means clustering analysis. Pearson statistical analysis of the three biofluids indicated that there was >90% similarity. Discovery DIA-MS analysis of the same samples using a plasma spectral library and a pan-human spectral library identified 1121 and 4661 total proteins, respectively. In addition, at least 122 FDA-approved biomarkers were identified. DIA-MS analysis reproducibly quantitated (<30% CV) ∼600-700 proteins in capillary blood, ∼800 proteins in venous blood, and ∼300-400 proteins in plasma, demonstrating that an expansive biomarker panel is possible with current mass spectrometry technology. Both targeted LC/MRM-MS and discovery DIA-MS analysis of whole blood collected on remote sampling devices are viable options for personal proteome biosignature stratification in precision medicine and precision health.

Evaluation of Small Molecules in Blank EDTA Plasma Tubes and Optimization of Metabolomic Workflow for Biomarker Studies Using Plasma Samples.

As the first step of metabolomic analysis in biomarker identification studies, various types of blood collection tubes are used in clinical practice. However, little attention is paid to potential contamination caused by the blank tube itself. Here, we evaluated small molecules in blank EDTA plasma tubes through LC-MS-based untargeted metabolomic analysis and identified small molecules with markedly varied levels among different production batches or specifications. Our data demonstrate possible contamination and data interference caused by blank EDTA plasma tubes when employing large clinical cohorts for biomarker identification. Therefore, we propose a workflow of filtering metabolites in blank tubes prior to statistical analysis to improve the fidelity of biomarker identification.

Automated and robust extraction of genomic DNA from various leftover blood samples.

With the development of genomic technologies, the isolation of genomic DNA (gDNA) from clinical samples is increasingly required for clinical diagnostics and research studies. In this study, we explored the potential of utilizing various leftover blood samples obtained from routine clinical tests as a viable source of gDNA. Using an automated method with optimized pre-treatments, we obtained gDNA from seven types of clinical leftover blood, with average yields of gDNA ranging from 3.11 ± 0.45 to 22.45 ± 4.83 µg. Additionally, we investigated the impact of storage conditions on gDNA recovery, resulting in yields of 8.62-68.08 µg when extracting gDNA from EDTA leftover blood samples stored at 4 °C for up to 13 weeks or -80 °C for up to 78 weeks. Furthermore, we successfully obtained sequenceable gDNA from both Serum Separator Tube and EDTA Tube using a 96-well format extraction, with yields ranging from 0.61 to 71.29 µg and 3.94-215.98 µg, respectively. Our findings demonstrate the feasibility of using automated high-throughput platforms for gDNA extraction from various clinical leftover blood samples with the proper pre-treatments.