OPTIMIZATION OF DRIED BLOOD SPOT ASSAYS FOR HEPATITIS B VIRUS SURFACE ANTIBODY QUANTIFICATION.

Dried blood spot (DBS) cards can be used as an alternative sample collection method to plasma, however, there is no optimized elution protocol for DBS cards specifically for hepatitis B surface antibody (anti-HBs) testing. The study aimed to develop a DBS elution protocol for anti-HBs quantification. Our study sought to determine the ideal phosphate-buffered saline (PBS) buffer volume to use by comparing three PBS volumes (300uL, 450uL, and 500uL), and the optimal time to agitate DBS discs on a plate shaker (1hr, 2hrs, 3hrs, and 4hrs) to yield DBS anti-HBs concentrations that are comparable to corresponding plasma anti-HBs concentrations. The optimal DBS storage temperature (25°C, -20°C, and -80°C) was investigated to determine the ideal long-term storage temperature of the cards. Residual samples were used for optimization (2019-2021). A total of 50 DBS-plasma pairs was used throughout the study, with plasma anti-HBs concentrations being used as the golden standard to compare. The analysis of results was carried out by determining the p-values of the Wilcoxon sign rank. A two-way analysis of variance (ANOVA) was also performed to determine the impact of PBS elution volumes, elution time, and storage temperature on the anti-HBs concentration of DBS samples on STATA Version 15.0. No statistically significant difference between the DBS-plasma anti-HBs pairs was observed when using 450 or 500uL of PBS buffer and when samples were agitated for 3 hours (p=0.594, p=0.499 respectively). The optimal storage temperature for DBS cards was 25°C because the results showed no statistically significant difference between DBS-plasma anti-HBs titers (p=0.594). The two-way ANOVA analysis showed that elution volumes and time had no statistically significant impact on the DBS anti-HBs concentrations, p=0.948 and p=0.381 respectively. Storage temperature had a statistically significant impact on the DBS anti-HBs concentrations, p=0.002. The optimized DBS elution protocol for anti-HBs quantification will help monitor vaccine efficacy in infants due to the low sample volumes required compared to plasma and also can be used for anti-HBs testing in resource-limited areas around the country.

Detection of doping substances in paired dried blood spots and urine samples collected during doping controls in Danish fitness centers.

The use of dried blood spot (DBS) in anti-doping can be advantageous in terms of collection, transportation, and storage compared with the traditional anti-doping testing matrices urine and venous blood. There could, nonetheless, be disadvantages such as shorter detection windows for some substances compared with urine, but real-life comparison of the detectability of prohibited substances in DBS and urine is lacking. Herein, we present a liquid chromatography-high resolution mass spectrometry (LC-HRMS)-based screening method for simultaneous detection of 19 target analytes from the doping substance categories S1-S5 in a single spot. Ninety-eight urine and upper-arm DBS (Tasso-M20) sample pairs were collected from fitness centers customers notified for doping control by Anti Doping Denmark, and three sample pairs were collected from active steroid users undergoing clinical evaluation and treatment at a Danish hospital. The analytical findings were cross compared to evaluate the applicability of the developed DBS testing menu in terms of feasibility and analytical performance. To our knowledge, this is the first study to compare the detectability of prohibited substances in DBS and urine samples collected in a doping control setting. Twenty-seven of the urine samples and 23 DBS samples were positive, and we observed a very high concordance (95%) in the overall analytical results (i.e., positive or negative samples for both urine and DBS). Collectively, these results are very promising, and DBS seems suitable as a stand-alone matrix in doping control in fitness centers likely because of the high analyte concentration levels in these samples.

Beyond the Tropics: Unraveling the Complex Relationship between Sun Exposure, Dietary Intake, and Vitamin D Deficiency in Coastal Malaysians.

Despite Malaysia's year-round sunny climate, vitamin D deficiency is surprisingly common among Malaysians. However, we hypothesise that vitamin D levels among coastal populations are above average. Thus, we aim to investigate vitamin D levels and correlate them with the potential contributing factors from three selected coastal villages in Johor, Melaka, and Negeri Sembilan. Convenient sampling was employed to recruit 120 Malay male and female participants, and dried blood spots (DBS) were obtained to measure 25 (OH) vitamin D3 levels via immunoassay. Participants also completed two questionnaires: the Sun Exposure and Protection Index (SEPI) and a validated food frequency questionnaire for Malaysians. The participant pool comprised 35.20% males and 64.80% females who completed all questionnaires and underwent DBS sampling. Our analysis revealed a significant difference (p < 0.05) based on skin tones, impacting various facets of the SEPI, including sunscreen usage, protective clothing utilisation, and the adoption of protective headwear. Furthermore, gender emerged as another pivotal factor, demonstrating significant distinctions in these SEPI components. Nevertheless, there is a weak correlation between SEPI scores and vitamin D levels. Subsequent regression analysis did produce statistically significant results (p = 0.018), yet the associated low R2 value indicated a weak correlation between dietary vitamin D intake that impacts vitamin D levels. In conclusion, our preliminary findings indicate that sun exposure and dietary factors are not the sole determinants of 25-OH vitamin D3 levels. However, we require more samples from various coastal locations for a definitive justification.

[Development of an analytical system for dried blood spots for forensic toxicology: a case study of five common drugs and poisons].

Dried blood spot (DBS) technology is a simple and convenient method for collecting, transporting, and storing blood samples on filter paper, and has numerous applications in the clinical, research, and public health settings. This technique is gaining popularity in the field of forensic science because it facilitates the rapid analysis of prohibited drugs in blood samples and offers significant advantages in toxicology scenarios such as drinking-driving screening, drug abuse detection, and doping detection. However, the lack of a standardized system and the fact that its stability and reliability have not been thoroughly researched and demonstrated limit its application in judicial practice in China. DBS samples can be prepared, stored, and analyzed in various ways, all of which may significantly affect the results. In this study, we developed a method based on ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) that focuses on the preparation, pretreatment, analysis, and storage of DBS samples. A thorough investigation was conducted to examine the optimal preparation conditions, including the blood spot matrix, drying technique, and preprocessing parameters, such as the solvent and extraction method. Moreover, the analytical conditions, such as the mobile phase system and elution gradient, were established to facilitate the quantitative detection of methamphetamine, lidocaine, ketamine, fentanyl, and diazepam in both DBS and whole-blood samples. The impact of storage conditions, such as the temperature, humidity, and sealing, on the analytical results of the DBS and whole-blood samples was also examined. The results showed a strong linear relationship for lidocaine and fentanyl within the range of 0.5-100 ng/mL. Similarly, methamphetamine, ketamine, and diazepam exhibited good linearity within the range of 2-100 ng/mL. The coefficients of determination (r2) ranged from 0.9983 to 0.9997, and the limits of detection ranged from 0.2 to 0.5 ng/mL, indicating a high degree of correlation and sensitivity. Stability tests demonstrated that the five target substances remained stable in the DBS for 60 days, with the measured contents deviating from the nominal values by 15%. Moreover, the measurement results of the DBS samples were highly similar to those of the whole-blood samples, with mean percentage differences of 4.44%, 3.50%, 7.66%, 5.10%, and 5.25% for fentanyl, diazepam, ketamine, lidocaine, and methamphetamine, respectively. Throughout the 60-day storage period, the maintenance of temperatures of -20 and 4 ℃, as well as sealing and dry storage, was not necessary. Room temperature was the most practical storage environment for the DBS samples. The results for each target showed very small concentration differences between the whole-blood and DBS samples, indicating that the DBS samples were suitable for drug and poison analysis in blood. Furthermore, the DBSs exhibited high quantitative consistency with the whole-blood samples, rendering them suitable matrices for preserving blood samples. Because DBS samples are easy to handle and store, they can realize the lightweight preservation of blood samples and provide a novel solution for the analysis and preservation of blood samples in public security practice. We recommend conducting comprehensive validations before utilizing DBS for analysis, particularly in terms of quantification, to ensure the judicial reliability of the results.

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.

Multiplex tandem mass spectrometry enzymatic activity assay for the screening and diagnosis of Mucolipidosis type II and III.

Mucolipidosis type II and III (MLII/III) is caused by defects in the mannose-6-phosphate system, which is essential to target most of the lysosomal hydrolases to the lysosome. MLII/III patients present with marked elevations in the activities of most lysosomal enzymes in plasma, but their profiles in dried blood spots (DBS) have not been well described. In the current study, we measured the activities of 12 lysosomal enzymes in DBS, among which acid sphingomyelinase, iduronate-2-sulfatase, and alpha-N-acetylglucosaminidase were significantly elevated in MLII/III patients when compared to random newborns. This sets the stage for using DBS to diagnose MLII/III. Furthermore, given an increasing number of lysosomal storage disorders are being included in the recommended uniform screening panel, our results also indicate that population-based newborn screening for MLII/III can be implemented with minimal efforts.

Importance to include differential diagnostics for acid sphingomyelinase deficiency (ASMD) in patients suspected to have to Gaucher disease.

The clinical manifestation of sphingolipidosis leads often to misclassification between acid sphingomyelinase deficiency (ASMD) and Gaucher disease. In this multicenter, prospective study, we investigated a cohort of 31,838 individuals suspected to have Gaucher disease, due to clinical presentation, from 61 countries between 2017 and 2022. For all samples, both Acid-ß-glucocerebrosidase and acid sphingomyelinase enzyme activities were measured in dried blood spot specimens by tandem mass spectrometry followed by genetic confirmatory testing in potential positive cases. In total, 5933 symptomatic cases showed decreased enzyme activities and were submitted for genetic confirmatory testing. 1411/5933 (24%) cases were finally identified with Gaucher disease and 550/5933 (9%) with ASMD. Most of the confirmed ASMD cases were newborns and children below 2 years of age (63%). This study reveals that one in four cases suspected for Gaucher disease is diagnosed with ASMD. An early appropriate diagnostic work-up is essential because of the availability of a recently approved enzyme replacement therapy for ASMD. In conclusion, a diagnostic strategy using differential biochemical testing including genetic confirmatory testing in clinically suspected cases for sphingolipidosis is highly recommended.

Precision sirolimus dosing in children: The potential for model-informed dosing and novel drug monitoring.

The mTOR inhibitor sirolimus is prescribed to treat children with varying diseases, ranging from vascular anomalies to sporadic lymphangioleiomyomatosis to transplantation (solid organ or hematopoietic cell). Precision dosing of sirolimus using therapeutic drug monitoring (TDM) of sirolimus concentrations in whole blood drawn at the trough (before the next dose) time-point is the current standard of care. For sirolimus, trough concentrations are only modestly correlated with the area under the curve, with R 2 values ranging from 0.52 to 0.84. Thus, it should not be surprising, even with the use of sirolimus TDM, that patients treated with sirolimus have variable pharmacokinetics, toxicity, and effectiveness. Model-informed precision dosing (MIPD) will be beneficial and should be implemented. The data do not suggest dried blood spots point-of-care sampling of sirolimus concentrations for precision dosing of sirolimus. Future research on precision dosing of sirolimus should focus on pharmacogenomic and pharmacometabolomic tools to predict sirolimus pharmacokinetics and wearables for point-of-care quantitation and MIPD.

Seroprevalence of SARS-CoV-2 antibodies in Italy in newborn dried blood spots.

OBEJCTIVES: Serosurveys can be used to monitor COVID-19 seroprevalence and conduct surveillance. Dried blood spot (DBS), used increasingly as a valuable sample to assay severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) antibodies (Ab), has several advantages, particularly in infants, due to the limited amount of blood required and its utility in testing a large number of samples in a limited time-frame. We evaluated SARS-CoV-2 IgG Ab prevalence in newborn DBS in the Trentino region of Italy, during the time period January 2020 - December 2021. METHODS: Anti-SARS-CoV-2 IgG levels were determined in DBS by means of Anti-SARS-CoV-2 QuantiVac IgG ELISA assay (Euroimmun, Lubeck, Germany). RESULTS: Analyses included 2,400 DBS from newborns (54% M, 46% F), samples being collected 2-3 days after birth. The first DBS that tested positive for anti-SARS-CoV-2 IgG antibodies was found in March 2020 and, up to May 2020, only 4 positive results were detected overall. Starting from June 2020, the positivity thresholds increased according to the epidemiological waves of the COVID-19 pandemic in Italy, with a robust increment in the winters of 2020 and 2021. The percentage of positive DBS rose from 0 to 6% to 10-47%, in 2020 and 2021, respectively. CONCLUSIONS: This study demonstrates DBS is a suitable tool for both epidemiological purposes and surveillance in the SARS-CoV-2 pandemic, particularly in newborns and pregnant women, saving blood waste and sparing patients any discomfort.

Diagnostic performance of the SARS-CoV-2 S1RBD IgG ELISA (ImmunoDiagnostics) for the quantitative detection of SARS-CoV-2 antibodies on dried blood spots.

Dried Blood Spots (DBS) are broadly used in SARS-CoV-2 surveillance studies, reporting either the presence or absence of SARS-CoV-2 antibodies. However, quantitative follow-up has become increasingly important to monitor humoral vaccine responses. Therefore, we aimed to evaluate the performance of DBS for the detection of anti-spike SARS-CoV-2 antibody concentrations using a commercially available assay, reporting in a standardised unitage (International Units/mL; IU/mL). To assess the sensitivity and specificity of the ImmunoDiagnostics ELISA on serum and DBS for SARS-CoV-2 antibody detection, we analysed 72 paired DBS and serum samples. The SARS-CoV-2 S1 IgG ELISA kit (EUROIMMUN) on serum was used as the reference method. We performed a statistical assessment to optimise the cut-off value for DBS and serum and assessed the correlation between DBS and serum antibody concentrations. We found that anti-spike SARS-CoV-2 antibody concentrations detected in DBS are highly correlated to those detected in paired serum (Pearson correlation 0.98; p-value < 0.0001), allowing to assess serum antibody concentration using DBS. The optimal cut-off for antibody detection on DBS was found to be 26 IU/mL, with 98.1% sensitivity and 100% specificity. For serum, the optimal cut-off was 14 IU/mL, with 100% sensitivity and 100% specificity. Therefore, we conclude that the ImmunoDiagnostics ELISA kit has optimal performance in the detection of SARS-CoV-2 antibodies on both DBS and serum. This makes DBS ideal for large-scale follow-up of humoral SARS-CoV-2 immune responses, as it is an easy but valuable sampling method for quantification of SARS-CoV-2 antibodies, compared to serum.

Non-Lethal Blood Sampling of Fish in the lab and Field With Methods for Dried Blood Plasma Spot Omic Analyses.

There is global acknowledgment that humane methods in animal research are a priority, but few environmental effects monitoring programs use nonlethal methods for fish. The goal of the present study was to determine the impacts of sampling small volumes of blood in larger-bodied fish on survival and healing. In addition to evaluating survival following blood sampling, we evaluated the utility of dried blood spots as an alternative for sample processing and storage in the field. In our approach, we housed 80 rainbow trout (Oncorhynchus mykiss) in our flow-through aquatic facility. We then anaesthetized using MS-222 and sampled 1 µl/g bw of blood via puncture of the caudal vasculature. We tested four different post-blood sampling treatments on the puncture wound: 1. application of liquid bandage; 2. a swab of betadine; 3. a swab of fish mucous; and 4. compared survival outcomes to a group where no post-treatment was performed (negative control). Overall, we observed 90% survival among all treatments, with the most effective approach being the negative control (100% survival). Based upon these results, we repeated the blood sampling with no-post treatment by housing 20 rainbow trout (not previously tested upon) in cages at a nearby creek and monitored survival for 2 weeks post sampling. The survival rate was 95% with full healing of the puncture site in all subjects. In addition to this, we tested the efficacy of dry blood spotting on proteomic, lipidomic and amino acid analysis as an alternative method for blood sample processing and storage. It was found that dried plasma spotting using parafilm in conjunction with a modified Bligh-Dyer extraction offered the best balance for good recovery of protein, lipid and amino acids relative to wet plasma and Noviplex dried plasma spot cards. In this article, we will present the detailed results of these combined studies and describe what we have determined to be the safest non-lethal blood sampling protocol.

Stability and detectability of testosterone esters in dried blood spots after intramuscular injections.

While misuse of testosterone esters is widespread in elite and recreational sports, direct detection of intact testosterone esters in doping control samples is hampered by the rapid hydrolysis by esterases present in the blood. With dried blood spot (DBS) as sample matrix, continued degradation of the esters is avoided due to inactivation of the hydrolase enzymes in dried blood. Here, we have developed the method further for detection of testosterone esters in DBS with focus on robustness and applicability in doping control. To demonstrate the method's feasibility, DBS samples from men receiving two intramuscular injections of Sustanon® 250 (n = 9) or placebo (n = 10) were collected, transported, and stored prior to analysis, to mimic a doping control scenario. The presented nanoLC-HRMS/MS method appeared reliable and suitable for direct detection of four testosterone esters (testosterone decanoate, isocaproate, phenylpropionate, and propionate) after extraction from DBS. Sustanon® was detected in all subjects for at least 5 days, with detection window up to 14 days for three of the esters. Evaluation of analyte stability showed that while storage at room temperature is tolerated well for a few days, testosterone esters are highly stable (>18 months) in DBS when stored in frozen conditions. Collectively, these findings demonstrate the applicability of DBS sampling in doping control for detection of steroid esters. The fast collection and reduced shipment costs of DBS compared with urine and standard blood samples, respectively, will allow more frequent and/or large-scale testing to increase detection and deterrence.

Comparison of conventional dried blood spots and volumetric absorptive microsampling for tacrolimus and mycophenolic acid determination.

Therapeutic drug monitoring (TDM) of immunosuppressants is essential to avoid either rejection or toxicity after solid organ transplantations. Capillary microsampling approaches are an outstanding alternative to conventional venous sampling for TDM (easy and non-invasive collection, enabling self-sampling, and cost-saving shipment, processing and storage). Volumetric absorptive microsampling (VAMS) has gained importance in the last years, as it was meant to overcome the hematocrit (Hct) related issues commonly associated to DBS analysis. Despite all the benefits, microsampling techniques performance (including a thorough clinical validation) should be set up before their implementation in clinical practice. The aim of this study was to perform a clinical validation for both tacrolimus (TAC) and mycophenolic acid (MPA) in both DBS and Mitra™ VAMS. For the clinical validations, two different requirements were set up: analytical (following EMA and FDA guidelines) and clinical (following the Royal College of Pathologists of Australasia -RCPA- recommendations) acceptance criteria. For DBS, both analytical and clinical acceptance criteria were fulfilled for TAC, with 98.7% and 95% of the paired samples within the preset limits, respectively. For MPA, the analytical criterion was met (70.6% of paired specimens), although only half of the pairs were within the clinical limits. For VAMS, the clinical validation for both TAC and MPA showed good correlations but significant lower concentrations in VAMS compared to the routine matrices. After VAMS concentrations correction, the analytical requirement was fulfilled for both analytes (71.1% for TAC, 75% for MPA), although the more restrictive criteria recommended by the RCPA were not met for any analyte (half of the samples fell within the acceptance area). In addition, no significant Hct impact on the quantification was found in any case. Also, a preliminary home-sampling trial was set up, showing promising results. Moreover, a comparison between VAMS vs. DBS analytical and clinical performances was carried out, including a home-sampling trial, sample quality results and costs. Although the analytical performance for both VAMS and DBS was similar, DBS were superior regarding clinical criteria, sampling quality and cost.

HIV-1 diagnosis using dried blood spots from patients in Kinshasa, DRC: a tool to detect misdiagnosis and achieve World Health Organization 2030 targets.

INTRODUCTION: Currently, only 54% of the population of the Democratic Republic of the Congo (DRC) know their HIV status. The aim of this study was to detect HIV misdiagnosis from rapid diagnostic tests (RDT) and to evaluate serological immunoassays using dried blood spots (DBS) from patients in Kinshasa, DRC. METHODS: Between 2016 and 2018, 365 DBS samples were collected from 363 individuals and shipped to Spain. The samples were from people with a new HIV positive (n = 123) or indeterminate (n = 23) result, known HIV-positive patients (n = 157), and a negative control group (n = 62). HIV serology was performed using Elecsys HIV combi PT (Roche), VIDAS HIV Duo Quick (BioMérieux), and Geenius (Bio-Rad). In addition, HIV RNA detection was performed in all samples using the COBAS AmpliPrep/COBAS Taqman HIV-1 Test 2.0 (Roche). RESULTS: Overall, 272 samples were found to be positive and 93 to be negative for HIV serology. The sensitivity was 100% for both Elecsys and VIDAS techniques, but specificity was slightly higher for the VIDAS test: 100% (96.1-100%) vs 98.9% (94.1-99.9%). Of the 23 indeterminate cases using RDT, only three cases were true-positives with a detectable viral load. Eleven samples out of the 280 classified as positive by RDT corresponded to nine patients who had received a false diagnosis of HIV through RDT (3.9%); six of them had been on antiretroviral therapy for at least 2 years. CONCLUSIONS: Elecsys HIV combi PT and VIDAS HIV Duo Quick immunoassays showed high sensitivity and specificity when using DBS. RDT-based serological diagnosis can lead to HIV misdiagnosis with personal and social consequences in sub-Saharan Africa.

No pain, just gain: Painless, easy, and fast dried blood spot collection from fingertip and upper arm in doping control.

This study aimed to determine and compare the perception, painfulness, and usability of the minimally invasive dried blood spot (DBS) collections from fingertip versus upper arm from different athlete populations: males and females representing sports dependent on hand/arm, sports less dependent on hand/arm and para-athletes. To accomplish this, 108 national level athletes from Denmark were recruited (♀ = 49, ♂ = 59, 25 ± 6 years; mean ± SD) and 11 Doping Control Officers (DCOs) collected manual fingerprick DBS (HemaSpot HF) and automated upper-arm DBS (Tasso-M20) from each athlete. Athletes and DCOs responded to questionnaires regarding the perception of sample collection procedures. On a 0-10 scale, the athletes reported a low pain score and a very good general experience for both sampling sites, but following upper-arm DBS collection, the associated pain was rated lower (-0.4 ± 1.6, p < 0.05), and the general experience rated better (+0.6 ± 2.3, p ≤ 0.001) than after the fingerprick DBS collection. The DCOs rated the general experience with the upper-arm DBS collection better (+1.6 ± 1.1, p ≤ 0.01) than the fingerprick DBS collection, partly because problems occurred more frequently during the DBS collection from the fingertip (28%) than from the upper arm (6%). In conclusion, it appears that DBS sampling is affiliated with minimal sensation of pain and is preferred by both DCOs and athletes, independent of gender and discipline, over conventional sample collection methods. Collection of DBS from the upper arm was preferred over fingerprick by both athletes and DCOs.

Expanded Screening of One Million Swedish Babies with R4S and CLIR for Post-Analytical Evaluation of Data.

Sweden has one neonatal screening laboratory, receiving 115 to 120 thousand samples per year. Among the one million babies screened by tandem mass spectrometry from November 2010 until July 2019, a total of 665 babies were recalled and 311 verified as having one of the diseases screened for with this methodology, giving a positive predictive value (PPV) of 47% and an incidence of 1:3200. The PPV was high (41%) already in the first year after start of screening, thanks to the availability of the collaborative project Region 4 Stork database. The PPV is presently 58%. This improvement was achieved by the implementation of second-tier analyses in the screening for methylmalonic aciduria, propionic aciduria, isovaleric aciduria, and homocystinuria, and the employment of various post analytical tools of the Region 4 Stork, and its successor the collaborative laboratory integrated reports.

Single-dose administration of clenbuterol is detectable in dried blood spots.

Clenbuterol is a ß2 -agonist prescribed for asthmatic patients in some countries. Based on its anabolic and lipolytic effects observed in studies on rodents and in livestock destined for food production, clenbuterol is abused by bodybuilders and athletes seeking leanness. Urinary clenbuterol analysis is part of routine doping analysis. However, the collection of urine samples is time-consuming and can be intimidating for athletes. Dried blood spot (DBS) appears attractive as an alternative matrix, but the detectability of clenbuterol in humans through DBS has not been investigated. This study evaluated if clenbuterol could be detected in DBS and urine collected from six healthy men after oral intake of 80 µg clenbuterol. The DBS and urine samples were collected at 0, 3, 8, 24, and 72 h post-ingestion, with additional urine collections on days 7 and 10. Using LC-MS/MS, it was shown that clenbuterol could be detected in all DBS samples for 24 h post-ingestion and with 50% sensitivity 3 days after ingestion. The DBS method was 100% specific. Evaluation of analyte stability showed that clenbuterol is stable in DBS for at least 365 days at room temperature when using desiccant and avoiding light exposure. In urine, clenbuterol was detectable for at least 7-10 days after ingestion. Urinary clenbuterol concentrations below 5 ng/mL were present in some subjects 24 h after administration. Collectively, these data indicate that DBS are suitable for routine doping control analysis of clenbuterol with a detection window of at least 3 days after oral administration of 80 µg.

Evaluation of the Xpert HCV VL Fingerstick point-of-care assay and dried blood spot HCV-RNA testing as simplified diagnostic strategies among people who inject drugs in Catalonia, Spain.

BACKGROUND: Catalonia requires decentralized and simplified strategies for the diagnosis of viremic HCV infection among people who inject drugs (PWID). We aimed to perform a direct comparison of the diagnostic performance between two, single-step strategies for the screening and diagnosis of viremic HCV infection in PWID attending a drug consumption room (DCR) in Barcelona: i) on-site HCV-RNA testing using the point-of-care test (PoCT) Xpert HCV VL Fingerstick; and ii) on-site dried blood spots (DBS) collection for HCV-RNA testing at the laboratory (in-house assay). Additionally, we aimed to assess participants' preferences in receiving HCV-RNA testing results and feasibility of same-day delivery of PoCT results. METHODS: The real-world, clinical performance of these two strategies was established in comparison with the reference method (HCV viral load testing with the Xpert HCV Viral Load assay at the laboratory from venous plasma collected at the DCR). HCV genotypes/subtypes and HIV status were also determined by sequencing and serology, respectively. A questionnaire including preferences regarding the delivery of test results was administered. RESULTS: The prevalence of HCV-RNA was 63.0% (of which 25.8% were co-infected with HIV). The RNA-PoCT showed a sensitivity of 98.4% for detectable viral loads (>4 IU/mL) and of 100% for quantifiable viral loads (≥10, ≥1000 and ≥3000 IU/mL). For the DBS-based assay, a sensitivity of 93.7% was obtained for detectable viral loads, 96.7% for the quantifiable ≥10 IU/mL threshold, and 98.3% for both the quantifiable ≥1000 and ≥3000 IU/mL thresholds. No significant differences were detected between the sensitivity values of these two strategies, and the specificity was 100% in both cases. Half of the participants preferred to receive the HCV-RNA result on the same day, and 80% of participants received their RNA-PoCT results on the same day. CONCLUSIONS: Both the HCV-RNA PoCT and the DBS-based assay are highly reliable tools for the simplified diagnosis of viremic HCV infection among current PWID. These strategies allow for on-site sample collection and delivery of test results, facilitating decentralized care in harm reduction services.

Fully automated dried blood spot sample preparation enables the detection of lower molecular mass peptide and non-peptide doping agents by means of LC-HRMS.

The added value of dried blood spot (DBS) samples complementing the information obtained from commonly routine doping control matrices is continuously increasing in sports drug testing. In this project, a robotic-assisted non-destructive hematocrit measurement from dried blood spots by near-infrared spectroscopy followed by a fully automated sample preparation including strong cation exchange solid-phase extraction and evaporation enabled the detection of 46 lower molecular mass (< 2 kDa) peptide and non-peptide drugs and drug candidates by means of LC-HRMS. The target analytes included, amongst others, agonists of the gonadotropin-releasing hormone receptor, the ghrelin receptor, the human growth hormone receptor, and the antidiuretic hormone receptor. Furthermore, several glycine derivatives of growth hormone-releasing peptides (GHRPs), arguably designed to undermine current anti-doping testing approaches, were implemented to the presented detection method. The initial testing assay was validated according to the World Anti-Doping Agency guidelines with estimated LODs between 0.5 and 20 ng/mL. As a proof of concept, authentic post-administration specimens containing GHRP-2 and GHRP-6 were successfully analyzed. Furthermore, DBS obtained from a sampling device operating with microneedles for blood collection from the upper arm were analyzed and the matrix was cross-validated for selected parameters. The introduction of the hematocrit measurement method can be of great value for doping analysis as it allows for quantitative DBS applications by managing the well-recognized "hematocrit effect." Graphical abstract.

Dried Blood Spot in Laboratory: Directions and Prospects.

Over the past few years, dried blood spot (DBS) technology has become a convenient tool in both qualitative and quantitative biological analysis. DBS technology consists of a membrane carrier (MC) on the surface of which a biomaterial sample becomes absorbed. Modern analytical, immunological or genomic methods can be employed for analysis after drying the sample. DBS has been described as the most appropriate method for biomaterial sampling due to specific associated inherent advantages, including the small volumes of biomaterials required, the absence of a need for special conditions for samples' storage and transportation, improved stability of analytes and reduced risk of infection resulting from contaminated samples. This review illustrates information on the current state of DBS technology, which can be useful and helpful for biomedical researchers. The prospects of using this technology to assess the metabolomic profile, assessment, diagnosis of communicable diseases are demonstrated.