Relay-race RNA/barcode gold nanoflower hybrid for wide and sensitive detection of microRNA in total patient serum

Development of a very sensitive biosensor is accompanied with an inevitable shrinkage in the linear detection range. Here, we developed an electrochemical biosensor with a novel methodology to detect microRNA-21 (miR21) at an ultralow level and broad linear detection range. A three-way junction RNA structure was designed harboring (i) a methylene blue (MB)-modified hairpin structure at its one leg to function as the sensing moiety and (ii) the other two legs to be further hybridized with barcode gold nanoparticles (MB/barG) as the signal amplifiers. Addition of target miR21 resulted in opening the hairpin moiety and subsequent hybridization with DNA-modified gold nanoflower/platinum electrode (GNF@Pt) to form the MB-3 sensor. Inspired by the relay-race run, to extend the dynamic detection range and increase the sensitivity of the biosensor, MB/barG was added to form the second detection modality (MBG-3). The combined sensor required very low sample volume (4 μL) and could identify 135 aM or 324 molecules of miR21 with the ability to operate within a wide linear range from 1 μM down to 500 aM. The fabricated GNF@Pt showed a remarkable conductivity compared with the gold nanoparticle-modified electrode. Addition of MB/barG boosted the electrochemical signal of the MB by almost 230 times. Moreover, a new protocol was introduced by the authors to increase the efficiency of microRNA extraction from the total serum. Possessing a sound selectivity and specificity towards single base-pair mutations, the developed biosensor could profile cancer development stages of two patient serums.