Phage-mediated counting by the naked eye of miRNA molecules at attomolar concentrations in a Petri dish.
The ability to count biomolecules such as cancer-biomarker miRNAs with the naked eye is seemingly impossible in molecular diagnostics. Here, we show an ultrasensitive naked-eye-counting strategy for quantifying miRNAs by employing T7 phage-a bacteria-specific virus nanoparticle-as a surrogate. The phage is genetically engineered to become fluorescent and capable of binding a miRNA-capturing gold nanoparticle (GNP) in a one-to-one manner. Target miRNAs crosslink the resultant phage-GNP couple and miRNA-capturing magnetic microparticles, forming a sandwich complex containing equimolar phage and miRNA. The phage is then released from the complex and developed into one macroscopic fluorescent plaque in a Petri dish by plating it in a host bacterial medium. Counting the plaques by the naked eye enables the quantification of miRNAs with detection limits of ∼3 and ∼5 aM for single-target and two-target miRNAs, respectively. This approach offers ultrasensitive and convenient quantification of disease biomarkers by the naked eye.
State Key Laboratory of Bioelectronics, Jiangsu Key Laboratory for Biomaterials and Devices, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, China.
Xin
X
Zhou
Laboratory of Cellular and Molecular Biology, Jiangsu Branch of China Academy of Chinese Medical Sciences, Nanjing 210028, China.
Peng
P
Cao
Department of Chemistry and Biochemistry, Stephenson Life Sciences Research Center, University of Oklahoma, 101 Stephenson Parkway, Norman, Oklahoma 73019-5300, USA.
Ye
Y
Zhu
Laboratory of Cellular and Molecular Biology, Jiangsu Branch of China Academy of Chinese Medical Sciences, Nanjing 210028, China.
Wuguang
W
Lu
State Key Laboratory of Bioelectronics, Jiangsu Key Laboratory for Biomaterials and Devices, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, China.
Ning
N
Gu
Department of Chemistry and Biochemistry, Stephenson Life Sciences Research Center, University of Oklahoma, 101 Stephenson Parkway, Norman, Oklahoma 73019-5300, USA.|School of Materials Science and Engineering, Zhejiang University, Hangzhou, Zhejiang 310027, China.
Chuanbin
C
Mao
D001419:Bacteria / Q000378:metabolism; D017123:Bacteriophage T7 / Q000737:chemistry; D001435:Bacteriophages / Q000502:physiology*; D015415:Biomarkers / Q000378:metabolism; D014408:Biomarkers, Tumor / Q000378:metabolism; D004247:DNA / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry; D016680:Genome, Bacterial; D006046:Gold / Q000737:chemistry; D006801:Humans; D053768:Metal Nanoparticles / Q000737:chemistry; D035683:MicroRNAs / Q000378:metabolism*; D046529:Microscopy, Electron, Transmission; D036103:Nanotechnology; D009369:Neoplasms; D009841:Oligonucleotides / Q000737:chemistry; D016133:Polymerase Chain Reaction; D015203:Reproducibility of Results
D015415:Biomarkers; D014408:Biomarkers, Tumor; D005456:Fluorescent Dyes; D035683:MicroRNAs; D009841:Oligonucleotides; D006046:Gold; D004247:DNA