Electrochemical immunosensor for N6-methyladenosine detection in human cell lines based on biotin-streptavidin system and silver-SiO2 signal amplification

N6-methyladenosine (m6A), a kind of RNA methylation form and important epigenetic event, plays crucial roles in many biological progresses. Thus it is essential to quantitatively detect m6A in complicated biological samples. Herein, a simple and sensitive electrochemical method was developed for m6A detection using N6-methyladenosine-5′-triphosphate (m6ATP) as detection target molecule. In this detection strategy, anti-m6A antibody was selected as m6A recognition and capture reagent, silver nanoparticles and amine-PEG3-biotin functionalized SiO2 nanospheres (Ag@SiO2) was prepared and used as signal amplification label, and phos-tag-biotin played a vital role of “bridge” to link m6ATP and Ag@SiO2 through the two forms of specific interaction between phosphate group of m6ATP and phos-tag, biotin and streptavidin, respectively. Under the optimal experimental conditions, the immunosensor presented a wide linear range from 0.2 to 500 nM and a low detection limit of 0.078 nM (S/N=3). The reproducibility and specificity were acceptable. Moreover, the developed method was also validated for detect m6A content in human cell lines. Importantly, this detection strategy provides a promising immunodetection platform for ribonucleotides and deoxyribonucleotides with the advantages of simplicity, low-costing, specificity and sensitivity.