An electrochemical biosensor based on DNAzyme and TB-loading mesoporous silica nanoparticles for detection of TdT in serum

Terminal deoxynucleotidyl transferase (TdT) is actively expressed in leukemia patients and plays an important role in maintaining the genetic diversity of an organism. Therefore, developing simple and sensitive analytical methods for the accurate detection of TdT in serum is of significant importance. Herein, an electrochemical biosensor based on a toluidine blue (TB)-loading mesoporous silica nanoparticles (MSN) was established for the detection of TdT in serum. First, MSN with amino group was synthesized and subsequently was carbonylated to obtain carboxyl-functionalized MSN (MSN-COOH), which was used to load TB. Thereafter, MSN-COOH was sealed with the substrate strand S(rA)-NH 2 of the Mg 2+ -dependent DNAzyme to form S(rA)-NH 2 /TB/MSN-COOH. In the presence of TdT, it can add deoxyadenosine triphosphate (dATP) to the 3’-OH end of Enc-5 A, activating its cleavage activity on the substrate chain S(rA)-NH 2 , thus releasing TB encapsulated in MSN-COOH. By measuring the change in the electrochemical signal of TB, the TdT concentration can be determined. The linear range is from 5 to 400 U/mL and the limit of detection (LOD) is 0.369 U/mL (3 σ/K). Since DNAzyme can be extended by TdT without the need of a template, the proposed biosensor demonstrates excellent selectivity towards TdT in the presence of other DNAzyme, such as T4 DNA ligase, phi29 DNA polymerase, DreamTaq DNA polymerase, and Klenow fragment ( exo -). Furthermore, the proposed biosensor was successfully applied to the detection of TdT in serum with the recovery rates ranged from 91.0 % to 94.7 %, indicating its potential for early detection of leukemia and other related diseases.