Electrochemical immunosensor for N6-methyladenosine detection in human cell lines based on biotin-streptavidin system and silver-SiO2signal 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 SiO2nanospheres (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@SiO2through 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 500nM and a low detection limit of 0.078nM (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.
College of Chemistry and Material Science, Shandong Agricultural University, Taian 271018, PR China.
Huanshun
H
Yin
College of Chemistry and Material Science, Shandong Agricultural University, Taian 271018, PR China.
Haiyan
H
Wang
College of Chemistry and Material Science, Shandong Agricultural University, Taian 271018, PR China.
Wenjing
W
Jiang
College of Chemistry and Material Science, Shandong Agricultural University, Taian 271018, PR China. Electronic address: zhyl@sdau.edu.cn.
Yunlei
Y
Zhou
College of Chemistry and Material Science, Shandong Agricultural University, Taian 271018, PR China. Electronic address: ashy@sdau.edu.cn.
Shiyun
S
Ai
D000241:Adenosine / Q000031:analogs & derivatives / Q000737:chemistry / Q000302:isolation & purification*; D015374:Biosensing Techniques; D002460:Cell Line; D055664:Electrochemical Techniques; D006046:Gold / Q000737:chemistry; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D053758:Nanoparticles / Q000737:chemistry*; D012834:Silver / Q000737:chemistry
D012834:Silver; D006046:Gold; D000241:Adenosine
Ag@SiO(2) nanocomposite; Electrochemical immunosensor; N6-methyladenosine; Phos-tag-biotin; RNA methylation