A Universal, Label‐Free, and Sensitive Optical Enzyme‐Sensing System for Nuclease and Methyltransferase Activity Based on Light Scattering of Carbon Nanotubes

Abstract A label‐free, enzyme‐responsive nanosystem that uses a DNA/single‐walled carbon nanotube (SWNT) assembly as the substrate is demonstrated for the sensitive, universal detection of restriction and nonrestriction endonucleases as well as methyltransferases in a homogeneous solution on the basis of light scattering (LS) of carbon nanotubes. This protocol is based on the different binding affinities of SWNTs to single‐ and double‐stranded DNA. This difference can lead to different LS signals that can be used for the detection of nuclease cleavage activity. The assay only requires a label‐free oligonucleotide probe, significantly reducing the typical cost. The LS technique and the use of a nuclease‐specific oligonucleotide probe impart extraordinarily high sensitivity and selectivity. This light scattering assay is universal and label‐free with a detection limit of 5 × 10 −6 U μL −1 for S1 nuclease, 1 × 10 −4 U μL −1 for EcoRI endonuclease, and 1 × 10 −2 U μL −1 for EcoRI methylase. In principle, this assay can be used to detect any kind of nuclease by simply changing the DNA sequences of the specific probe.