DNAzymes-conjugated upconversion nanoamplicon for in-situ ultrasensitive detection and imaging of microRNA in vivo

• A microRNA upconversion cycling nanoamplicon sensing platform was fabricated. • 8-17 DNAzymes were employed as nanoamplicon. • The nanoprobe showed high LRET efficiency of 60.9% for BHQ1 as the energy acceptor. • The nanoprobe achieved a linear range from 4×10 -14 M to 1×10 -8 M with LOD of 31 fM. • The nanoprobe was successfully applied to in - vitro and in - vivo imaging of miR-21. Upconversion nanoparticles (UCNPs) are promising materials for imaging biomolecules and predicting the progression of diseases, especially for tumors. However, conventional luminescence resonance energy transfer (LRET) based nanosensors remain challenging with complicated modification and unsatisfactory sensitivity. Herein, a sensing platform integrating confined emission effect and cycling amplification strategy into one substrate with high LRET efficiency and sensitivity has been developed for the in - vitro and in - vivo detection of microRNA-21. Washing UCNPs with hydrochloric acid was conducted for electrostatic absorption of 8-17 DNAzyme labeled with BHQ1 (8-17 DNAzyme-BHQ1) and complementary capturing template-aDNA3 complex (CPT-aDNA3) to form energy donor-acceptor pair (UCNPs-BHQ1) and capture miR-21. It not only avoids the complex modification procedure but also shortens the distance between the donor (UCNPs) and the acceptor (BHQ1). The quenching efficiency of 60.94% was obtained. Targeted miR-21 could bind with CPT to release aDNA3 that can unlock 8-17 DNAzyme-BHQ1 to form aDNA3-DNAzyme-BHQ1 duplex. This aDNA3- DNAzyme-BHQ1 duplex owning -rA- group can be cleaved by Zn 2+ so that UCNPs restored green UCL emission because of the desorption of BHQ1. aDNA3 becomes dissociative again to trigger the next cycle. This UCNP@DNAzyme-BHQ1/CPT- aDNA3 achieved a wide linear range from 4×10 -14 M to 1×10 -8 M with a low limit detection of 31 fM for the sensitive detection of miR-21 in vitro . Furthermore, it was successfully applied to the real-time intracellular detection of miR-21 in HeLa cells and in - vivo monitoring of miR-21 expression in KS-01 treated tumor-bearing mouse. This proposed DNA-conjugated upconversion nanoamplicon provides new sights into monitoring of anti-cancer active ingredients efficacy and the progress evaluation of miRNA-related diseases.