New Technology Could Improve Detection of Biomarkers for CF, Other Diseases

A new strategy for detecting very small concentrations of specific molecules could allow for better diagnosis and management of a variety of diseases, including cystic fibrosis (CF).

The method was described in the journal Nature Communication by a team from the University of Leeds, U.K., in a study titled “Rational design of DNA nanostructures for single molecule biosensing.”

Measuring the amount of molecules in the blood or other bodily tissue can provide important information related to health — for example, levels of C-reactive protein (CRP) in the blood are associated with inflammation. CRP measurements are important for the management of many conditions, including CF.

Measuring the levels of certain molecules requires the use of specialized technology. For instance, enzyme-linked immunosorbent assays (ELISAs) are a common method for measuring concentrations that uses antibodies to detect a molecular target.

However, there are inherent limitations as to how sensitively these technologies can detect their targets — an ELISA to detect CRP requires a certain minimum amount of the protein to be present, just to generate a detectable signal.

Developing technologies that can more precisely perform these measurements is an ongoing effort in biomedical research — the “ideal” technology would, in theory, be able to detect single molecules in a sample.

“The ability to detect biomarkers with single entity resolution rather than via ensemble-averaging techniques provides significant advantages for the detection of ultra-small biomarker concentrations,” the researchers wrote.

The Leeds team therefore developed a new technique based on two concepts: nanopores and DNA origami.

Nanopores refer to very small holes. Specific adaptations can be designed so that the nanopore is a sensor for a range of specific molecules.

As its name suggests, DNA origami involves taking a strand of DNA and “folding” it to create a specific structure. These structures can be tailored such that they have a central cavity that will only bind to a specific molecular target.

In the study, the researchers basically used DNA origami to create nanopores that could only be “filled” by a single, specific molecule.

“The captured biomarkers [target molecules] are then read with nanopores and we can do this one molecule at a time,” Mukhil Raveendran, the study’s first author, said in a press release. “By coupling DNA origami and nanopores we are able to quantitatively detect disease biomarkers with single molecule sensitivity.”

As a proof of concept, the researchers developed an an analysis based on these principles to measure CRP. They showed that this assay could detect small CRP concentrations in serum — as low as 9 nanomolar.

Importantly, the assay did not detect MupB, a protein that is about the same size as CRP, suggesting the assay is specific for CRP identification.

“One of the main advantages [of the technique] is the minimal sample needed,” Raveendran said. “We are able to isolate individual molecules from small samples to identify specific illnesses. The process is very quick, and takes just minutes to provide results.”

The researchers are now working to adapt the technology to detect other molecular targets of interest, including molecules from the virus that causes COVID-19.

“Sensitive detection of biomarkers is important for diagnosis and for disease management,” said Paolo Actis, PhD, a professor at Leeds and the study’s senior author. “We have already demonstrated the detection of an inflammation marker called C-reactive protein (important for the management of many diseases including cystic fibrosis) in diluted serum.”