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Fluorogenic assays for activated protein C using aptamer modified magnetic beads |
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| Authors: | Qiang Zhao Jie Gao |
| Affiliation: | 1. Research Center for Environmental Science and Engineering, Shanxi University, Taiyuan, 030006, China 2. College of Chemistry and Chemical Engineering, Shanxi University, Taiyuan, 030006, China |
| Abstract: | We describe a fluorogenic assay for activated protein C (APC) by using magnetic beads modified with DNA aptamers, taking advantage of strong binding affinity of aptamer, facile magnetic separation, and signal amplification via an enzymatic reaction. APC is specifically captured from a sample by the DNA aptamers on magnetic beads, and the concentrated APC then catalyzes the conversion of a fluorogenic substrate of APC to a fluorescent product. Detection of APC is achieved by measuring the generated product. This method is simple, sensitive, and specific. APC can be detected at 0.4 pM concentration level in a sample volume of 250 μL, corresponding to 0.1 femtomole of APC, when 2-h enzymatic reaction is employed. The proteins thrombin, trypsin, proteinase K, chymotrypsin, and elastase do not interfere. An aptamer-based assay for human activated protein C (APC) is developed. APC is specifically captured from sample mixture by the aptamers on magnetic beads, and the captured APC converts substrate to product. Measurement of the generated product allows the determination of APC. |
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