A triple signal amplification method for chemiluminescent detection of the cancer marker microRNA-21

Mesoporous silica nanospheres (MSNs) are used in a triple signal amplification chemiluminescent (CL) assay for microRNA-21. It is based on (a) the synergistic amplification via loading and controlled-release of signal reagents by MSNs, (b) target recycling amplification, and (c) the enhancement effect of graphene oxide quantum dots (GOQD). CL is generated by the bis(2,4,6-trichlorophenyl) oxalate (TCPO) and H2O2 reaction in the presence of the fluorophore rhodamine B (RB). RB is firstly loaded into the pores of MSNs modified with amino groupsand coupled with ssDNA. Then, the pores are capped by GOQD. Upon the addition of microRNA-21 into the system, the designed ssDNA assumes a double stranded structure. With the aid of duplex-specific nuclease, the double strand structure is cleaved and the free microRNA-21 enters into the next cycling process to combine with other ssDNA forming double strand structures. After several cycling process, amounts of GOQDs departing from the surface of MSNs cause the opening of the pores of MSNs and the release of RB causes the CL of TCPO-H2O2 reaction system. Free GOQDs can lead to a further CL enhancement. By this method, even a low amount of microRNA-21 leads to a large number of released RB molecules and triggers high-intensity CL. The method was applied in an assay where the CL signal increases linearly with the logarithm of the microRNA-21 concentration in the range of 0.005–50 pmol L−1 and the detection limit is 1.7 fmol L−1 (at 3σ).