Journal
ANALYTICAL CHEMISTRY
Volume 83, Issue 24, Pages 9234-9236Publisher
AMER CHEMICAL SOC
DOI: 10.1021/ac203063z
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Funding
- Natural Sciences and Engineering Research Council of Canada
- Canadian Institutes of Health Research
- Canada Research Chairs Program
- Alberta Health and Wellness
- National Natural Science Foundation of China [20905043]
- Shanxi Scholarship Council of China
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Activity and specificity of enzyme molecules are important to enzymatic reactions and enzyme assays. We describe an aptamer capturing approach that improves the specificity and the sensitivity of enzyme detection. An aptamer recognizing the target enzyme molecule is conjugated on a magnetic bead, increasing the local concentration, and serves as an affinity probe to capture and separate minute amounts of the enzyme. The captured enzymes catalyze the subsequent conversion of fluorogenic substrate to fluorescent products, enabling a sensitive measure of the active enzyme. The feasibility of this technique is demonstrated through assays for human alpha thrombin and human neutrophil elastase (HNE), two important enzymes. Thrombin (2 fM) and 100 fM HNE can be detected. The incorporation of two binding events, substrate recognition and aptamer binding, greatly improves assay specificity. With its simplicity, this approach is applicable to biosensing and detection of disease biomarkers.
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