Detection of Individual Proteins Bound along DNA Using Solid-State Nanopores

Area of Science:

• Nanotechnology
• Molecular Biology
• Biophysics

Background:

• Cellular DNA is regulated by numerous proteins, making DNA-bound protein detection a significant challenge.
• Solid-state nanopores offer a linear readout of DNA and associated protein volumes within a molecular constriction.

Purpose of the Study:

• To demonstrate the detection of individual DNA-bound proteins at the single-DNA-molecule level using solid-state nanopores.
• To establish a novel model system using anti-DNA antibodies and lambda phage DNA for nanopore detection.

Main Methods:

• Utilized solid-state nanopores for high-resolution translocation measurements of DNA molecules.
• Employed a model system of anti-DNA antibodies bound to lambda phage DNA, chosen for individual binding and stability in high salt concentrations.
• Analyzed translocation events for characteristic current changes indicative of bound antibodies.

Main Results:

• Successfully detected individual DNA-bound antibodies at the single-DNA-molecule level.
• Observed short, distinct current spikes (12 microseconds) during translocation, with amplitudes approximately 4.5 times larger than that of double-stranded DNA (dsDNA).
• Attributed these signals to the transient interactions between the antibodies and the nanopore during translocation.

Conclusions:

• Solid-state nanopores can effectively detect individual DNA-bound proteins.
• The developed model system provides a robust platform for studying DNA-protein interactions.
• This research serves as a proof-of-concept for future nanopore-based biosensing applications.