scholarly journals A surrogate virus neutralization test to quantify antibody-mediated inhibition of SARS-CoV-2 in finger stick dried blood spot samples

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Amelia E. Sancilio ◽  
Richard T. D’Aquila ◽  
Elizabeth M. McNally ◽  
Matthew P. Velez ◽  
Michael G. Ison ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Venous Blood ◽  
Host Cells ◽  
Dried Blood Spot ◽  
Blood Collection ◽  
Widespread Application ◽  
Live Virus ◽  
Dilution Series ◽  
Wide Range ◽  
Blood Spot

AbstractThe spike protein of SARS-CoV-2 engages the human angiotensin-converting enzyme 2 (ACE2) receptor to enter host cells, and neutralizing antibodies are effective at blocking this interaction to prevent infection. Widespread application of this important marker of protective immunity is limited by logistical and technical challenges associated with live virus methods and venous blood collection. To address this gap, we validated an immunoassay-based method for quantifying neutralization of the spike-ACE2 interaction in a single drop of capillary whole blood, collected on filter paper as a dried blood spot (DBS) sample. Samples are eluted overnight and incubated in the presence of spike antigen and ACE2 in a 96-well solid phase plate. Competitive immunoassay with electrochemiluminescent label is used to quantify neutralizing activity. The following measures of assay performance were evaluated: dilution series of confirmed positive and negative samples, agreement with results from matched DBS-serum samples, analysis of results from DBS samples with known COVID-19 status, and precision (intra-assay percent coefficient of variation; %CV) and reliability (inter-assay; %CV). Dilution series produced the expected pattern of dose–response. Agreement between results from serum and DBS samples was high, with concordance correlation = 0.991. Analysis of three control samples across the measurement range indicated acceptable levels of precision and reliability. Median % surrogate neutralization was 46.9 for PCR confirmed convalescent COVID-19 samples and 0.1 for negative samples. Large-scale testing is important for quantifying neutralizing antibodies that can provide protection against COVID-19 in order to estimate the level of immunity in the general population. DBS provides a minimally-invasive, low cost alternative to venous blood collection, and this scalable immunoassay-based method for quantifying inhibition of the spike-ACE2 interaction can be used as a surrogate for virus-based assays to expand testing across a wide range of settings and populations.

scholarly journals A surrogate virus neutralization test to quantify antibody-mediated inhibition of SARS-CoV-2 in finger stick dried blood spot samples

2021 ◽  
Author(s):  
Amelia Sancilio ◽  
Richard D’Aquila ◽  
Elizabeth M. McNally ◽  
Matt E Velez ◽  
Michael G. Ison ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Venous Blood ◽  
Host Cells ◽  
Dried Blood Spot ◽  
Blood Collection ◽  
Widespread Application ◽  
Live Virus ◽  
Dilution Series ◽  
Wide Range ◽  
Blood Spot

ABSTRACTBackgroundThe spike protein of SARS-CoV-2 engages the human angiotensin-converting enzyme 2 (ACE2) receptor to enter host cells, and neutralizing antibodies are effective at blocking this interaction to prevent infection. Widespread application of this important marker of protective immunity is limited by logistical and technical challenges associated with live virus methods and venous blood collection. To address this gap, we validated an immunoassay-based method for quantifying neutralization of the spike-ACE2 interaction in a single drop of capillary whole blood, collected on filter paper as a dried blood spot (DBS) sample.MethodsSamples are eluted overnight and incubated in the presence of spike antigen and ACE2 in a 96-well solid phase plate. Competitive immunoassay with electrochemiluminescent label is used to quantify neutralizing activity. The following measures of assay performance were evaluated: dilution series of confirmed positive and negative samples, agreement with results from matched DBS-serum samples, analysis of results from DBS samples with known COVID-19 status, and precision (intra-assay percent coefficient of variation; %CV) and reliability (inter-assay; %CV).ResultsDilution series produced the expected pattern of dose-response. Agreement between results from serum and DBS samples was high, with concordance correlation = 0.991. Analysis of three control samples across the measurement range indicated acceptable levels of precision and reliability. Median % neutralization was 46.9 for PCR confirmed convalescent COVID-19 samples and 0.1 for negative samples.ConclusionsLarge-scale testing is important for quantifying neutralizing antibodies that can provide protection against COVID-19 in order to estimate the level of immunity in the general population. DBS provides a minimally-invasive, low cost alternative to venous blood collection, and this scalable immunoassay-based method for quantifying neutralization of the spike-ACE2 interaction can be used as a surrogate for virus-based assays to expand testing across a wide range of settings and populations.

scholarly journals Validation of dried blood spot sample modifications to two commercially available COVID-19 IgG antibody immunoassays

Bioanalysis ◽  
2020 ◽  
Author(s):  
Theodore T Zava ◽  
David T Zava
Keyword(s):  
Venous Blood ◽  
Dried Blood Spot ◽  
Blood Collection ◽  
Igg Antibodies ◽  
Antibody Testing ◽  
Igg Antibody ◽  
Nucleocapsid Proteins ◽  
Antibody Assays ◽  
Finger Stick ◽  
Blood Spot

Aim: Coronavirus disease 2019 antibody testing often relies on venous blood collection, which is labor-intensive, inconvenient and expensive compared with finger-stick capillary dried blood spot (DBS) collection. The purpose of our work was to determine if two commercially available anti-severe acute respiratory syndrome coronavirus 2 enzyme-linked immunosorbent assays for IgG antibodies against spike S1 subunit and nucleocapsid proteins could be validated for use with DBS. Materials & methods: Kit supplied reagents were used to extract DBS, and in-house DBS calibrators were included on every run. Results: Positive/negative concordance between DBS and serum was 100/99.3% for the spike S1 subunit assay and 100/98% for the nucleocapsid assay. Conclusion: Validation of the DBS Coronavirus disease 2019 IgG antibody assays demonstrated that serum and DBS can produce equivalent results with minimal kit modifications.

scholarly journals Enzyme immunoassay for SARS-CoV-2 antibodies in dried blood spot samples: A minimally-invasive approach to facilitate community- and population-based screening

2020 ◽  
Author(s):  
Thomas W. McDade ◽  
Elizabeth M. McNally ◽  
Richard D’Aquila ◽  
Brian Mustanski ◽  
Aaron Miller ◽  
...  
Keyword(s):  
Minimally Invasive ◽  
Venous Blood ◽  
Population Based ◽  
Dried Blood Spot ◽  
Molecular Diagnostic ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serological Testing ◽  
Igg Antibodies ◽  
Wide Range ◽  
Blood Spot

AbstractBackgroundSerological testing for SARS-CoV-2 IgG antibodies is needed to document the community prevalence and distribution of the virus, particularly since many individuals have mild symptoms and cannot access molecular diagnostic testing of naso-pharyngeal swabs. However, the requirement for serum/plasma limits serological testing to clinical settings where it is feasible to collect and process venous blood. To address this problem we developed a serological test for SARS-CoV-2 IgG antibodies that requires only a single drop of capillary whole blood, collected from a simple finger prick and dried on filter paper (dried blood spot, DBS).MethodsEnzyme linked immunosorbent assay (ELISA) was optimized to detect SARS-CoV-2 IgG antibodies against the receptor-binding domain (RBD) of the spike protein. DBS samples were eluted overnight and transferred to a 96-well plate coated with antigen, and anti-human IgG-HRP was used to generate signal in proportion to bound antibody. DBS samples spiked with anti-SARS IgG antibody, and samples from known positive and negative cases, were compared to evaluate assay performance.ResultsAnalysis of samples with known concentrations of anti-SARS IgG produced the expected pattern of dose-response. Optical density (OD) values were significantly elevated for known positive cases in comparison with samples from unexposed individuals.DiscussionDBS ELISA provides a minimally-invasive alternative to venous blood collection that combines the convenience of sample collection in the home or non-clinical setting with the quantitation of ELISA in the lab. Serological testing for SARS-CoV-2 IgG antibodies in DBS samples should facilitate research across a wide range of community- and population-based settings on seroprevalence, predictors and duration of antibody responses, as well as correlates of protection from reinfection, each of which is critically important for pandemic control.

scholarly journals Evaluation of Dried Blood Spot Sampling for Clinical Metabolomics: Effects of Different Papers and Sample Storage Stability

Metabolites ◽  
2019 ◽  
Vol 9 (11) ◽  
pp. 277 ◽  
Author(s):  
Trifonova ◽  
Maslov ◽  
Balashova ◽  
Lokhov
Keyword(s):  
Glass Fiber ◽  
Storage Stability ◽  
Direct Injection ◽  
Venous Blood ◽  
Ion Source ◽  
Dried Blood Spot ◽  
Blood Collection ◽  
Sample Storage ◽  
Sample Collection ◽  
Blood Spot

The dried blood spot (DBS) sampling has a lot of advantages in comparison with the “standard” venous blood collecting, such as small collection volume, painless and easy sample collection with minimal training required, stable and transportable at ambient temperatures, etc. The aim of this study was to determine the comparability of four different types of DBS sampling (HemaSpot™-HF Blood Collection Device, Whatman® 903 Protein Saver Snap Apart Card, card ImmunoHealth™, and glass fiber strip ImmunoHealth™) for analysis of the global metabolites profile. All the samples were collected from the same person at the same time and stored at room temperature for four weeks in order to exclude all possible deviations deriving from biological variances and to evaluate sample storage stability. Metabolome profiling by direct injection of a deproteinized capillary blood DBS sample into an electrospray ion source of a hybrid quadrupole time-of-flight mass spectrometer was used. Differences in the metabolomics profile were found between the different DBS collection materials, especially for ImmunoHealth™ card and ImmunoHealth™ glass fiber strip. However, our results indicate that the analytical performance of all tested DBS sampling materials showed consistent results overall detected metabolites and no dramatic changes between them in the metabolic composition during the storage time.

scholarly journals Dried Blood Spot Analysis Suitable for Therapeutic Drug Monitoring of Voriconazole, Fluconazole, and Posaconazole

2013 ◽  
Vol 57 (10) ◽  
pp. 4999-5004 ◽  
Author(s):  
Kim C. M. van der Elst ◽  
Lambert F. R. Span ◽  
Kai van Hateren ◽  
Karin M. Vermeulen ◽  
Tjip S. van der Werf ◽  
...  
Keyword(s):  
Therapeutic Drug Monitoring ◽  
Drug Monitoring ◽  
Sampling Method ◽  
Fungal Infections ◽  
Venous Blood ◽  
Blood Sampling ◽  
Dried Blood Spot ◽  
Therapeutic Drug ◽  
Venous Blood Sampling ◽  
Blood Spot

ABSTRACTInvasive aspergillosis and candidemia are important causes of morbidity and mortality in immunocompromised and critically ill patients. The triazoles voriconazole, fluconazole, and posaconazole are widely used for the treatment and prophylaxis of these fungal infections. Due to the variability of the pharmacokinetics of the triazoles among and within individual patients, therapeutic drug monitoring is important for optimizing the efficacy and safety of antifungal treatment. A dried blood spot (DBS) analysis was developed and was clinically validated for voriconazole, fluconazole, and posaconazole in 28 patients. Furthermore, a questionnaire was administered to evaluate the patients' opinions of the sampling method. The DBS analytical method showed linearity over the concentration range measured for all triazoles. Results for accuracy and precision were within accepted ranges; samples were stable at room temperature for at least 12 days; and different hematocrit values and blood spot volumes had no significant influence. The ratio of the drug concentration in DBS samples to that in plasma was 1.0 for voriconazole and fluconazole and 0.9 for posaconazole. Sixty percent of the patients preferred DBS analysis as a sampling method; 15% preferred venous blood sampling; and 25% had no preferred method. There was significantly less perception of pain with the DBS sampling method (P= 0.021). In conclusion, DBS analysis is a reliable alternative to venous blood sampling and can be used for therapeutic drug monitoring of voriconazole, fluconazole, and posaconazole. Patients were satisfied with DBS sampling and had less pain than with venous sampling. Most patients preferred DBS sampling to venous blood sampling.

PKP-014 Dried blood spot sampling of nilotinib in patients with chronic myeloid leukaemia: a comparison with venous blood samples

2014 ◽  
Vol 21 (Suppl 1) ◽  
pp. A142.1-A142
Author(s):  
AM Schimmel ◽  
CCLM Boons ◽  
A Chahbouni ◽  
AJ Wilhelm ◽  
YM Den Hartog ◽  
...  
Keyword(s):  
Chronic Myeloid Leukaemia ◽  
Myeloid Leukaemia ◽  
Venous Blood ◽  
Dried Blood Spot ◽  
Blood Samples ◽  
Blood Spot

scholarly journals Penicillin Dried Blood Spot Assay for Use in Patients Receiving Intramuscular Benzathine Penicillin G and Other Penicillin Preparations To Prevent Rheumatic Fever

2017 ◽  
Vol 61 (8) ◽  
Author(s):  
Madhu Page-Sharp ◽  
Jonathan Coward ◽  
Brioni R. Moore ◽  
Sam Salman ◽  
Lewis Marshall ◽  
...  
Keyword(s):  
Rheumatic Fever ◽  
Venous Blood ◽  
Plasma Concentrations ◽  
Management Plan ◽  
Dried Blood Spot ◽  
Penicillin G ◽  
Poor Correlation ◽  
Benzathine Penicillin ◽  
Limit Of Quantification ◽  
Blood Spot

ABSTRACT Rheumatic heart disease (RHD) remains an important global health challenge. Administration of benzathine penicillin (BPG) every 3 to 4 weeks is recommended as a secondary prophylaxis to prevent recurrent episodes of acute rheumatic fever and subsequent RHD. Following intramuscular injection, BPG is hydrolyzed to penicillin G (benzylpenicillin). However, little is known of the pharmacokinetics (PK) of BPG in pediatric populations at high risk of RHD or of the pharmacokinetic-pharmacodynamic relationship between penicillin exposure and clinically relevant outcomes. Dried blood spot (DBS) assays can facilitate PK studies in situations where frequent venous blood sampling is logistically difficult. A liquid chromatography-mass spectroscopy assay for penicillin G in plasma and DBS was developed and validated. Application of the DBS assay for PK studies was confirmed using samples from adult patients receiving penicillin as part of an infection management plan. The limit of quantification for penicillin G in DBS was 0.005 mg/liter. Penicillin G is stable in DBS for approximately 12 h at room temperature (22°C), 6 days at 4°C, and >1 month at −20°C. Plasma and DBS penicillin G concentrations for patients receiving BPG and penicillin G given via bolus doses correlated well and had comparable time-concentration profiles. There was poor correlation for patients receiving penicillin via continuous infusions, perhaps as a result of the presence of residual penicillin in the peripherally inserted central catheter, from which the plasma samples were collected. The present DBS penicillin G assay can be used as a surrogate for plasma concentrations to provide valid PK data for studies of BPG and other penicillin preparations developed to prevent rheumatic fever and RHD.

scholarly journals Receptor Binding, Fusion Inhibition, and Induction of Cross-Reactive Neutralizing Antibodies by a Soluble G Glycoprotein of Hendra Virus

2005 ◽  
Vol 79 (11) ◽  
pp. 6690-6702 ◽  
Author(s):  
Katharine N. Bossart ◽  
Gary Crameri ◽  
Antony S. Dimitrov ◽  
Bruce A. Mungall ◽  
Yan-Ru Feng ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
Gradient Centrifugation ◽  
Hendra Virus ◽  
Host Cells ◽  
Cell Surface Receptor ◽  
Size Exclusion ◽  
Emerging Viruses ◽  
Fusion Event ◽  
Live Virus

ABSTRACT Hendra virus (HeV) and Nipah virus (NiV) are closely related emerging viruses comprising the Henipavirus genus of the Paramyxovirinae, which are distinguished by their ability to cause fatal disease in both animal and human hosts. These viruses infect cells by a pH-independent membrane fusion event mediated by their attachment (G) and fusion (F) glycoproteins. Previously, we reported on HeV- and NiV-mediated fusion activities and detailed their host-cell tropism characteristics. These studies also suggested that a common cell surface receptor, which could be destroyed by protease, was utilized by both viruses. To further characterize the G glycoprotein and its unknown receptor, soluble forms of HeV G (sG) were constructed by replacing its cytoplasmic tail and transmembrane domains with an immunoglobulin κ leader sequence coupled to either an S-peptide tag (sGS-tag) or myc-epitope tag (sGmyc-tag) to facilitate purification and detection. Expression of sG was verified in cell lysates and culture supernatants by specific affinity precipitation. Analysis of sG by size exclusion chromatography and sucrose gradient centrifugation demonstrated tetrameric, dimeric, and monomeric species, with the majority of the sG released as a disulfide-linked dimer. Immunofluorescence staining revealed that sG specifically bound to HeV and NiV infection-permissive cells but not to a nonpermissive HeLa cell line clone, suggesting that it binds to virus receptor on host cells. Preincubation of host cells with sG resulted in dose-dependent inhibition of both HeV and NiV cell fusion as well as infection by live virus. Taken together, these data indicate that sG retains important native structural features, and we further demonstrate that administration of sG to rabbits can elicit a potent cross-reactive neutralizing antibody response against infectious HeV and NiV. This HeV sG glycoprotein will be exceedingly useful for structural studies, receptor identification strategies, and vaccine development goals for these important emerging viral agents.

scholarly journals Optimization of extraction of genomic DNA from archived dried blood spot (DBS): potential application in epidemiological research & bio banking

2018 ◽  
Vol 2 ◽  
pp. 57 ◽  
Author(s):  
Abhinendra Kumar ◽  
Sharayu Mhatre ◽  
Sheela Godbole ◽  
Prabhat Jha ◽  
Rajesh Dikshit
Keyword(s):  
Genomic Dna ◽  
Large Scale ◽  
Venous Blood ◽  
Magnetic Bead ◽  
Epidemiological Studies ◽  
Dried Blood Spot ◽  
Future Research ◽  
Prolonged Storage ◽  
Whole Genome ◽  
Blood Spot

Background: Limited infrastructure is available to collect, store and transport venous blood in field epidemiological studies. Dried blood spot (DBS) is a robust potential alternative sample source for epidemiological studies & bio banking. A stable source of genomic DNA (gDNA) is required for long term storage in bio bank for its downstream applications. Our objective is to optimize the methods of gDNA extraction from stored DBS and with the aim of revealing its utility in large scale epidemiological studies.  Methods: The purpose of this study was to extract the maximum amount of gDNA from DBS on Whatman 903 protein saver card. gDNA was extracted through column  (Qiagen) & magnetic bead based (Invitrogen) methods. Quantification of extracted gDNA was performed with a spectrophotometer, fluorometer, and integrity analyzed by agarose gel electrophoresis.  Result: Large variation was observed in quantity & purity (260/280 ratio, 1.8-2.9) of the extracted gDNA. The intact gDNA bands on the electrophoresis gel reflect the robustness of DBS for gDNA even after prolonged storage time. The extracted gDNA amount 2.16 – 24 ng/µl is sufficient for its PCR based downstream application, but unfortunately it can’t be used for whole genome sequencing or genotyping from extracted gDNA. Sequencing or genotyping can be achieved by after increasing template copy number through whole genome amplification of extracted gDNA. The obtained results create a base for future research to develop high-throughput research and extraction methods from blood samples. Conclusion: The above results reveal, DBS can be utilized as a potential and robust sample source for bio banking in field epidemiological studies.

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