Label-free photosensitization colorimetric detection of microRNA by utilizing cascade toehold assembly amplification

The sensitive and label-free detection of microRNA (miRNA) is crucial for the comprehension of miRNA-related pathological process, such as glioma. Here, we utilized a SYBR Green I (SG) catalytic photosensitization colorimetric reaction in conjunction with target recognition-induced exonuclease-iii (Exo-iii) assisted cascade toehold assembly amplification to achieve highly sensitive and label-free miRNA detection. In this research, the photosensitized generation of ¹O₂ by SG could directly oxidize 3,3′,5,5′-tetramethylbenzidine (TMB) without the need for H₂O₂ oxidation or enzyme catalysis, which simplifies the experimental procedures and renders the method highly stable. Furthermore, the Exo-iii enhances the target recognition induced catalytic chain assembly, thereby forming cascade signal amplification and providing the method with a significantly higher level of sensitivity than conventional colorimetric miRNA detection methods. Based on this, the proposed method exhibits a low limit of detection of 2.1 fM for miRNA-212 detection. Taking the merit of the high sensitivity, portability, and simplicity, the proposed method can serve as a highly effective nucleic acid detection tool and can facilitate the advancement of biosensors for point-of-care testing (POCT) and clinical disease diagnosis.