Highly Sensitive Electrochemiluminescence Assay for Cardiac Troponin I and Adenosine Triphosphate by using Supersandwich Amplification and Bifunctional Aptamer

Abstract

A highly sensitive electrochemiluminescence (ECL) assay was developed for sequential detection of cardiac troponin I (cTnI) and adenosine triphosphate (ATP) through supersandwich amplification and bifunctional aptamer. The bifunctional aptamer (S1) contained ATP binding aptamer and cTnI binding aptamer. A gold electrode was modified with the specific peptide of cTnI through a self-assembly technique. A sandwich-type conjugate (peptide S1) was formed when the peptide-modified electrode was successively reacted with cTnI and S1. Then, hybridizations between S1 and two ss-DNA auxiliary probes, in which one is complementary with ATP binding aptamer and the other is ATP binding aptamer, led to the formation of long-range ds-DNA on the electrode surface. In the presence of ECL indicator Ru(phen)32+, a large amount of Ru(phen)32+ was intercalated into ds-DNA grooves, resulting in amplification of the ECL signals. The ECL assay was successfully developed for the detection of cTnI in the range of 8.0×10−13 to 1.0×10−11 g/mL based on the increased ECL intensity. After detecting cTnI, ATP was detected in the range of 30 to 500 nM, based on switching structures of aptamers from ds-DNA to ss-DNA/target complex. A low detection limit of 0.3 pg/mL and 10 nM for cTnI and ATP, respectively, was obtained. The employment of a bifunctional aptamer probe and supersandwich signal amplification is promising for the sensitive detection of multiple targets.