(Nanowerk Spotlight) In modern medicine, the ability to detect disease markers early can mean the difference between life and death. Yet, detecting biomarkers like tumor necrosis factor-alpha (TNF-α) at extremely low levels, critical for early diagnosis, has long posed a challenge.
TNF-α acts as a signaling molecule in the body, facilitating communication between cells and influencing processes like inflammation, cell death, and immune response. The ability to detect TNF-α early and at very low concentrations can lead to earlier diagnoses and better treatment outcomes for patients suffering from diseases like rheumatoid arthritis and certain cancers. Yet despite advances in biomolecular detection technologies, current biosensing methods often struggle to detect TNF-α at the minuscule concentrations present in the early stages of disease.
Recent research into biosensing technologies has produced promising developments in this area. Dr. Shuwen Zeng and her team at L2n (light, nanomaterials, nanotechnologies) Laboratory, French National Centre for Scientific Research (CNRS), in collaboration with Prof. Megan Yi-Ping Ho from the Department of Biomedical Engineering, Chinese University of Hong Kong, have introduced a novel biosensing platform that combines aptamer-functionalized surface plasmon resonance (SPR) with an optical effect known as the Goos-Hänchen (GH) shift. The GH shift is a phenomenon where light, upon reflection, experiences a tiny lateral movement. By measuring this movement, the sensor can detect even the slightest binding of molecules, offering a higher sensitivity than traditional methods.
This development enables the detection of TNF-α at femtomolar concentrations (10-15 M), a sensitivity level previously unattainable without complex signal amplification strategies.
GH-aptasensing of TNF-α. (Image: Dr. Shuwen Zeng and Dr. Kathrine N. Borg)
Antibodies, the proteins often used in biosensors, are effective but come with drawbacks such as high cost and variability between batches. In contrast, the use of aptamers – a type of nucleic acid that binds specifically to its target – provides a more stable, cost-effective, and versatile alternative.
“The most significant result of our study is the integration of the Goos-Hänchen shift into aptamer-functionalized surface plasmon resonance biosensing, enabling the detection of TNF-α at femtomolar concentrations,” Dr. Zeng and Dr. Borg, the paper’s first author, explain. “This is a breakthrough as traditional aptamer-based SPR systems typically detect cytokines at higher detection limits, often in the nanomolar range. By utilizing aptamers as recognition units and coupling them with the ultrasensitive GH shift, we achieved a sensitivity level that surpasses most conventional optical sensors”
Surface plasmon resonance has long been a go-to technique in biosensing, allowing researchers to detect molecular interactions in real time by measuring changes in the refractive index at the surface of a sensor. However, detecting small molecules like TNF-α, which are present in very low concentrations, has posed a significant challenge.
The research team’s breakthrough came with the combination of SPR with the GH shift, a phenomenon that occurs when light is reflected at an interface, causing a slight lateral shift in the light beam. By precisely measuring this shift, the biosensor can detect even the smallest changes caused by biomolecule interactions, pushing the limits of sensitivity beyond what traditional SPR methods can achieve.
The platform’s aptamer-functionalized surface provides further advantages. Aptamers, selected through a process called SELEX (Systematic Evolution of Ligands by Exponential Enrichment) – a method for selecting nucleic acid sequences that bind to specific targets with high affinity – are synthetic single-stranded nucleic acids that bind specifically to a target, in this case, TNF-α. These aptamers are more stable than antibodies, easier to produce, and highly customizable, making them ideal for biosensing applications.
The team’s system immobilizes aptamers on the sensor’s surface, where they bind with TNF-α molecules flowing through a microfluidic system. This binding event induces a measurable GH shift, which signals the presence of the target biomolecule.
In explaining the broader impact of this development, Dr. Zeng noted, “The unprecedented sensitivity of the GH-aptasensing platform positions it as a pivotal tool for the early detection of diseases where TNF-α is a key biomarker, such as inflammatory disorders. Detecting low levels of TNF-α can lead to timely interventions, potentially improving patient prognosis. Beyond mere biosensing, this platform also opens avenues for in-depth exploration of cytokine-aptamer interactions, enabling researchers to investigate affinity kinetics and thermodynamics. Such insights can enhance our understanding of disease mechanisms and inform the development of targeted therapies”
One of the key findings of this research is the platform’s ability to detect TNF-α at a concentration as low as 1 femtomolar, a detection limit that surpasses many current technologies. For comparison, conventional aptamer-based SPR sensors typically operate in the nanomolar range, as covered in their review article (Advanced Optical Materials, “Recent Developments on Optical Aptasensors for the Detection of Pro-Inflammatory Cytokines with Advanced Nanostructures”), and even antibody-based immunoassays often require signal amplification to reach clinically relevant sensitivity levels
The ability to detect TNF-α at such low concentrations is critical for early diagnosis, especially in diseases where early detection can drastically alter treatment outcomes.
Highlighting the platform’s impact on diagnostics, Dr. Zeng points out that “this achievement not only establishes a new benchmark for cytokine detection but also holds promise for facilitating earlier diagnosis and significantly improving treatment strategies and patient outcomes. As such, this result represents a substantial advancement in clinical diagnostics with important implications for improving patient care.”
The versatility of the GH-aptasensing platform extends beyond TNF-α. The researchers plan to expand its capabilities to detect other biomarkers, such as interleukin-6 (IL-6), another significant marker in inflammatory diseases. Additionally, they are exploring the potential for multiplexed biosensing, which would allow for the simultaneous detection of multiple biomarkers in a single test. This capability could be especially useful in clinical settings, providing a more comprehensive picture of a patient’s condition by detecting a panel of disease-related biomarkers.
One of the advantages of using aptamers in biosensing is their adaptability. Aptamers can be designed to target a wide range of molecules, from proteins and peptides to small organic compounds and even cells. This flexibility, combined with the sensitivity of the GH shift, makes the platform an attractive candidate for point-of-care testing, where rapid and sensitive results are essential.
Dr. Zeng envisions broad applications for the platform: “Beyond healthcare, this technology could be adapted for detecting environmental pollutants or pathogens at trace levels, ensuring public health safety. The potential applications of this technology could significantly advance patient care and research in biomarker discovery.”
However, despite these promising developments, challenges remain. Translating the platform from laboratory settings to clinical use will require extensive validation, particularly in complex biological samples like blood, where nonspecific interactions could affect accuracy. The scalability and cost-effectiveness of producing these sensors for widespread clinical use also pose challenges. Addressing these issues will be key to realizing the full potential of this technology.
The team is already looking ahead to the next steps in their investigation. “While our current work has demonstrated excellent sensitivity for TNF-α, we plan to further explore binding kinetics and thermodynamics of both bovine serum albumin and TNF-α,” Dr. Zeng concludes. “This deeper understanding of the molecular interactions will help optimize the system for clinical applications. Additionally, we aim to conduct tests with spiked serum samples to assess the platform’s performance in real-world biological contexts, further validating its robustness and reliability.”
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