This study details trends in direct alcohol biomarker concentrations from civil cases within the United Kingdom (UK). Our subject cohort in this study related to family law litigation, where an individual was subject to an alcohol monitoring order by the court. This monitoring was conducted by quantification of alcohol biomarkers Phosphatidlyethanol (PEth) in dried blood spots (DBS) and Ethyl Glucuronide (EtG) and Ethyl Palmitate (EtPa) from hair segments. In total 298 PEth cases predominantly from the South East of England during the period July 2022 to August 2023 were analysed for alcohol biomarkers in DBS and hair. Subjects alcohol intake was classified as abstinence/low alcohol consumption, moderate or excessive alcohol consumption, based on a combination of Society for Hair Testing and PEth Net guidelines. Our results indicate that 33 % of PEth concentrations were consistent with excessive alcohol use (>200 ng/mL DBS), with 36 % consistent with social or moderate alcohol use (20-200 ng/mL DBS). In relation to EtG and EtPa 23 % and 31 % of subjects were classified as excessive alcohol users respectively. This study indicates that DBS sampling of PEth is a more sensitive predictor of alcohol use, in particular, at differentiating between moderate and excessive alcohol use compared to EtG and EtPa testing in hair. The authors suggest that increased frequency in the sampling of PEth in DBS (multiple occasions per month) may provide a more accurate assessment and simplification of the interpretation criteria of alcohol patterns rather than the combined hair testing and DBS sampling that are typically requested by UK courts.

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.

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.

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.

BACKGROUND: 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.
CONCLUSIONS: 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.

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.

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.

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-β-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.

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.

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.