An RNA Aptamer-Based Electrochemical Biosensor for Detection of Theophylline in Serum

Abstract

An electrochemical RNA aptamer-based biosensor for rapid and label-free detection of the bronchodilator theophylline was developed. The 5'-disulfide-functionalized end of the RNA aptamer sequence was immobilized on a gold electrode, and the 3'-amino-functionalized end was conjugated with a ferrocene (Fc) redox probe. Upon binding of theophylline the aptamer switches conformation from an open unfolded state to a closed hairpin-type conformation, resulting in the increased electron-transfer efficiency between Fc and the electrode. The electrochemical response, which was measured by differential pulse voltammetry, reaches saturation within a few minutes after addition of theophylline, and the dynamic range for detecting theophylline is 0.2-10 muM. The electrode displays an inhibited response when applied directly in serum samples treated with RNase inhibitors; however a full response to the theophylline serum concentration was obtained by transferring the electrode to blank serum-free buffer solutions. It was demonstrated that theophylline is detected with high selectivity in the presence of caffeine and theobromine.

Real-time measurement of small molecules directly in awake, ambulatory animals

“…endogenous metabolites and hormones in rat blood activates the sensor, as evidenced by their performance in vivo. The platform's ability to distinguish between structurally similar molecules, in contrast, can be problematic due to the sometimes [although not always (43,44)] limited specificity of aptamers because, of course, the sensor cannot be more specific than the aptamer from which it is constructed. E-AB specificity is nevertheless sufficient for many research and clinical applications.…”

Metal–Carbon Bonds in Biopolymer Conjugates: Bioorganometallic Nucleic Acid Chemistry

“…15 Amide bonds have also been used to attach a variety of other linkers containing ferrocene moieties to the 5¤- 16 and 3¤-ends of DNA 17,18 and RNA. 19 Other simple postsynthetic systems for attaching ferrocene to the ends of DNA strands include a nitrogen phosphate 2, 20 a carbodiimide group that reacts with the imino groups of thymines, uridines, and guanines 21 and an oxime bond forming reaction used to tag both the 3¤-and 5¤-ends of DNA 3.…”

Establishment of an electrochemical RNA aptamer-based biosensor to trace nanomolar concentrations of codeine

“…to complete the 3'-terminus reaction. The electrode was then rinsed with phosphate buffer (pH 7.0) gently and was treated with 2 mM 1-mercaptohexanol in phosphate buffer (pH 7.0) for 2 h. 19,24,25 By performing these procedures, the electrode was made ready and was immediately used in experiments.…”