Self-contained G-quadruplex/hemin DNAzyme: a superior ready-made catalyst for in situ imaging analysis.

The G4 DNAzyme holds significant potential for applications in bioanalysis and determination owing to its peroxidase mimetic activity and DNA programmability. However, its clinical practicability is constrained by limited catalytic activity and supplementary assembly requirements, attributed to weak π-π stacking, deficient active-site components, and ion-dependent assembly mechanisms. Thus, we constructed a highly active self-contained intramolecular G4/hemin DNAzyme through the direct covalent cross-linking of catalytic core components involving the hemin prosthetic group, G4 pocket, and distal ligand-like assistant nucleotide (adenine or cytosine). Detailed investigations of the catalytic efficiency and mechanism confirmed the formation of a compact catalytic active center through covalent bonding, which enhanced the catalysis to a stage comparable to that of horseradish peroxidase in localized surroundings. The superior ready-made catalytic modularity with programmability enabled the highly sensitive in situ imaging analysis of HER2 protein in breast cancer specimens. This study provides a powerful tool for disease marker imaging detection with high sensitivity and immediate availability.