Target catalyzed hairpin assembly for constructing a ratiometric electrochemical aptasensor.

In this paper, we develop a novel dual-signaling amplified aptasensor for protein detection via target-catalyzed hairpin assembly. Thrombin was chosen as a model target. This aptasensor contains two DNA hairpins termed as H1 and H2. H1, which is modified at its 3' ends with a methylene blue (MB), consists of the aptamer sequence of human thrombin. Meanwhile, H2 which is modified at its 3' ends with a ferrocene (Fc), is partially complementary to H1. Upon the addition of target protein, it can facilitate the opening of the hairpin structure of H1 and thus accelerate the hybridization between H1 and H2, the target protein can be displaced from hairpin H1 by hairpin H2 through a process similar to DNA branch migration. The released target found another H1 to trigger the cycle, resulting in the multiplication of the Fc confined near the GE surface and MB away from the GE surface. When IFc/IMB is used as the response signal for quantitative determination of thrombin, the detection limit (41 fM) is much lower than that by using either MB or Fc alone. This new dual-signaling aptasensor is readily regenerated and shows good response toward the target. Furthermore, this amplified aptasensor shows high selectivity toward its target protein. The clever combination of the functional DNA hairpin and the novel device achieved a ratiometric electrochemical aptasensor, which could be used as a simple, sensitive high repeatability and selective platform for target protein detection.