Applying CRISPR-Cas12a as a Signal Amplifier to Construct Biosensors for Non-DNA Targets in Ultralow Concentrations.
Junjie LiShuangshuang YangChen ZuoLing DaiYongcan GuoGuoming XiePublished in: ACS sensors (2020)
Efficient signal amplification is essential to construct ultrasensitive biosensors for biologically relevant species with abundant concomitant interferences. Here, we apply LbaCas12a as a signal amplifier to develop a versatile CRISPR-Cas12a platform to detect a wide range of analytes in ultralow concentrations. The platform relies on the indiscriminate single-stranded DNase activity of LbaCas12a, which recognizes single-stranded DNA intermediates generated by non-DNA targets down to femtomolar concentrations and subsequently enhances the fluorescence signal output. With the help of functional nucleotides (DNAzyme and aptamer), ultrasensitive bioassays for Pb2+ and Acinetobacter baumannii have been designed with a limit of detection down to ∼0.053 nM and ∼3 CFU/mL, respectively. It also allows simultaneous detection of four microRNAs (miRNAs) at a picomolar concentration without significant interferences by other counterparts, suggesting the potential of multiplexed miRNA expression profiles analysis in high throughput. Given the versatility and generality of the CRISPR-Cas12a platform, we expect the current work to advance the application of CRISPR-Cas-based platforms in bioanalysis and provide new insights into ultrasensitive biosensor design.
Keyphrases
- label free
- crispr cas
- high throughput
- genome editing
- acinetobacter baumannii
- single molecule
- circulating tumor
- nucleic acid
- cell free
- gold nanoparticles
- multidrug resistant
- drug resistant
- single cell
- pseudomonas aeruginosa
- binding protein
- quantum dots
- heavy metals
- photodynamic therapy
- circulating tumor cells
- energy transfer
- sensitive detection
- climate change
- living cells
- data analysis