A fully-automated, six-plex single molecule immunoassay for measuring cytokines in blood
| Autor: | Cheuk W. Kan, David C. Duffy, David M. Rissin, Todd G. Campbell, Derek R. DuPont, Andrew J. Rivnak, Carlos Cabrera, David Fournier, Sean Sullivan, Melissa Gardel, Brian A. Pink, Matthew W. Fishburn, Tomasz Piech, Linan Song |
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| Rok vydání: | 2015 |
| Předmět: |
Protein biomarkers
medicine.medical_treatment Immunology Enzyme-Linked Immunosorbent Assay Workflow Automation Crohn Disease Diabetes Mellitus medicine Humans Immunology and Allergy Multiplex Immunoassay Detection limit biology medicine.diagnostic_test Chemistry Reproducibility of Results Molecular biology Cytokine Fully automated biology.protein Cytokines Antibody After treatment |
| Zdroj: | Journal of Immunological Methods. 424:20-27 |
| ISSN: | 0022-1759 |
| Popis: | We report a system and assay for performing fully-automated measurement of 6 proteins simultaneously with single molecule sensitivity. The system combines handling of samples, reagents, and consumables, with a module for imaging single molecule arrays (Simoa) to enable immunoassays that have high sensitivity (~fg/mL), are multiplexed, and are fully-automated. A 6-plex cytokine Simoa assay for IL-6, TNF-α, GM-CSF, IL-10, IL-1β, and IL-1α was developed on the system. The assays had limits of detection in the range 0.01-0.03pg/mL, and the average imprecision (CV) of the Simoa signal was 4.2%. This assay was used to measure the concentrations of these cytokines in the plasma of patients with Crohn's Disease (CD), before and after treatment with anti-TNF-α antibody drugs, and in the serum of Type 1 diabetics. Concentrations of TNF-α and IL-6 in the CD samples determined using the fully-automated, multiplex Simoa assay had good correlation with the manual, single-plex assays previously reported. Drug treatment caused reductions in the mean concentration of all 6 cytokines in the plasma of CD patients. The concentrations of 4 cytokines were significantly higher in diabetics compared to healthy controls. The system could enable the widespread, multiplexed measurement of protein biomarkers with low abundance. |
| Databáze: | OpenAIRE |
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