## 3 Results and Discussion
### Sensitivity and Selectivity of the ECL Biosensor  
Because each target DNA molecule is recycled in TRA, the RCA reaction could theoretically generate numerous tandem repeats containing G-quadruplex sequences that result in a signal that is amplified thousands of times, and our proposed genosensor is expected to provide high sensitivity with amplification of the intercalated ECL signal quencher hemin in SaOs\({}^{2-}\) buffer. The ECL response to genosensors associated with different concentrations of target DNA in testing buffers serve as the basic value for target determination. As shown in Figure 5A, a relatively strong basic ECL response was obtained in the absence of target DNA. However, the addition of an increasing amount of target DNA resulted in a dynamic decrease in ECL intensity. Because more target DNA led to the generation of more hemin/G-quadruplex that could competitively consume the dissolved oxygen in detection buffer, there was a higher quenching effect toward the O\({}_{2}\)/S\({}_{2}\)Os\({}^{2-}\) system. This strategy demonstrates a good relationship between the concentration of target DNA and ECL intensity (y\(=-9371.2-980.1\) log C, C: concentration of target DNA (M), y: ECL intensity) with a good regression coefficient (\(R^{2}=0.99\)). The linear range was from 10.0 aM to 1.0 pM for the target DNA (Figure 5B), and the estimated detection limit was 3.8 aM (3\(\sigma\)). Moreover, the coefficient of variation (CV, \(n=6\)) of the target DNA at the 10.0 fM level with different electrodes from one batch was 6.5%, showing good intraassay reproducibility for the ECL sensor. Interassay reproducibility was also tested with six repetitive measurements at 10.0 fM target DNA with one electrode and a CV of 5.7% was obtained. Figure 5: Sensitivity of modified electrodes. (**A**) ECL responses of the sensors to different concentrations of target DNA: (a) 0 M; (b) 10.0 aM; (c) 100.0 aM; (d) 1.0 fM; (e) 10.0 fM; (f) 100.0 fM; (g) 1.0 pM; (**B**) The resulting calibration plot of log c _vs._ ECL intensity (y \(=-9371.2-980.1\) logC; C: concentration of target DNA(M), y: ECL intensity, \(R^{2}=0.99\)). ECL measurement conditions were performed as described in Figure 3B, and all experiments were repeated 3 times (error bars \(=\) SD, \(n=3\)). To evaluate the selectivity of the proposed genosensor, we compared the change in ECL intensity caused by the complementary (target), single-base mismatched (sDNA) and non-complementary sequence (nDNA). As shown in Figure 6A, the ECL signals for 100.0 fM sDNA (column c) and nDNA (column b)were slightly weaker than that of the control experiment (column a), which had an absence of target DNA. However, the presence of a lower (10-fold) concentration of target DNA (column d, 10.0 fM) yielded a significantly weaker ECL signal compared with the blank and excess nontarget DNA. This comparison suggests that only perfect match DNA triggers the TRA and RCA processes efficiently. Thus, the TRA and RCA system is capable of identifying complementary target DNA sequences and nontarget sequences, suggesting good selectivity for our proposed genosensor. Moreover, the stability of the proposed TRA and RCA-based DNA assay was also examined by scanning a continuous CV for 6 cycles (shown in Figure 6B), and accordingly, sharp ECL peaks with almost constant intensity was obtained, suggesting good stability of this sensor. The recovery experiments were performed by standard addition methods in human serum to monitor the feasibility of the developed ECL genosensor. The results were shown in Table 2 and the recovery was in the range of 96.1%-103.0%, which demonstrated that our strategy could be considered as an optional scheme for detection of _S. aureus_ DNA in clinical diagnostics.