Advancing Precision in DNA Analysis: Super-Resolution Technique Scans Individual Molecules

Researchers at EPFL, led by Dr. Aleksandra Radenovic, have made significant progress in nanopore technology by integrating it with scanning ion conductance microscopy. The resulting technique, known as scanning ion conductance spectroscopy (SICS), offers unparalleled precision in controlling the speed of molecular transit, resulting in a substantial increase in signal-to-noise ratio. This versatile method has the potential to revolutionize DNA analysis, proteomics, and clinical research. Credit: Samuel Leitão / EPFL

EPFL researchers have achieved remarkable control over the manipulation and characterization of individual molecules, achieving unprecedented precision.

Dr. Aleksandra Radenovic, the head of the Laboratory of Nanoscale Biology in the School of Engineering, has dedicated years of work to improving nanopore technology. This technology involves passing molecules, such as DNA, through a minuscule pore in a membrane to measure an ionic current. By analyzing how each nucleotide perturbs this current as it passes through, scientists can determine the DNA’s sequence of genetic information. The research, published in the journal Nature Nanotechnology on June 19, represents a significant breakthrough.

Currently, the passage of molecules through a nanopore and the timing of their analysis are influenced by random physical forces. The rapid movement of molecules poses challenges in achieving high analytical accuracy. Radenovic has previously addressed these issues using optical tweezers and viscous liquids. Now, a collaboration with Georg Fantner and his team in the Laboratory for Bio- and Nano-Instrumentation at EPFL has yielded the breakthrough she has been striving for, with implications that extend beyond DNA.

By combining nanopore technology with scanning ion conductance microscopy for the first time, EPFL researchers have achieved near-perfect control over the manipulation and identification of individual molecules, resulting in unprecedented precision. Credit: Samuel Leitão / EPFL

“We have merged the sensitivity of nanopores with the precision of scanning ion conductance microscopy (SICM), enabling us to target specific molecules and locations and control their movement speed. This exceptional control could bridge a significant gap in the field,” says Radenovic. The researchers accomplished this control by utilizing a state-of-the-art scanning ion conductance microscope, developed at the Lab for Bio- and Nano-Instrumentation.

Enhancing sensing precision by two orders of magnitude

The fortuitous collaboration between the two labs was sparked by PhD student Samuel Leitão. His research focuses on SICM, a technique that uses variations in the ionic current flowing through a probe tip to generate high-resolution 3D image data. For his PhD, Leitão developed and applied SICM technology to image nanoscale cell structures, employing a glass nanopore as the probe. In this recent work, the team harnessed the precision of a SICM probe to guide molecules through a nanopore, rather than relying on random diffusion.

Named scanning ion conductance spectroscopy (SICS), this innovation slows down the transit of molecules through the nanopore. This allows for thousands of consecutive readings to be taken on the same molecule, or even at different locations along the molecule. The ability to control the transit speed and obtain multiple readings of the same molecule has resulted in a two-order-of-magnitude increase in the signal-to-noise ratio compared to conventional methods.

By combining nanopore technology with scanning ion conductance microscopy for the first time, EPFL researchers have achieved near-perfect control over the manipulation and identification of individual molecules, resulting in unprecedented precision. Credit: Samuel Leitão / EPFL

“What is particularly exciting is that this enhanced detection capability with SICS could potentially be applied to other solid-state and biological nanopore methods, leading to significant improvements in diagnostic and sequencing applications,” adds Leitão.

Fantner summarizes the approach with an automotive analogy: “Imagine standing in front of a window, watching cars drive back and forth. It becomes much easier to read their license plate numbers if the cars slow down and drive by repeatedly,” he explains. “We also have the choice to measure 1,000 different molecules once or the same molecule 1,000 times, representing a true paradigm shift in the field.”

The precision and versatility of this approach open the door to its application with molecules beyond DNA, such as peptides, which are the building blocks of proteins. This advancement could have significant implications for proteomics, as well as biomedical and clinical research.

“Sequencing peptides has been a major challenge due to the complexity of their ‘license plates,’ which consist of 20 characters (amino acids), unlike DNA’s four nucleotides,” says Radenovic. “For me, the most exciting prospect is that this newfound control might pave the way for easier peptide sequencing.”

Reference: “Spatially multiplexed single-molecule translocations through a nanopore at controlled speeds” by S. M. Leitao, V. Navikas, H. Miljkovic, B. Drake, S. Marion, G. Pistoletti Blanchet, K. Chen, S. F. Mayer, U. F. Keyser, A. Kuhn, G. E. Fantner and A. Radenovic, 19 June 2023, Nature Nanotechnology.
DOI: 10.1038/s41565-023-01412-4

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• EPFL News: Super-Resolution DNA Analysis
• Nature Nanotechnology: Spatially multiplexed single-molecule translocations through a nanopore at controlled speeds