As part of an extensive effort to radically change how pain is diagnosed, researchers have created an innovative microfluidic biosensor that has been termed the “pain-on-a-chip.” The sensor is designed to identify pain signatures from small blood samples and could provide a pivotal shift in how chronic pain is monitored and addressed from an objective, biological standpoint. Chronic pain is a global health issue that affects millions of people and imposes a significant burden on quality of life.
Pain is one of the most common medical complaints and a major reason for prolonged disability. Pain can be categorized as either acute, which typically lasts three months or less for recovery, or chronic, which persists beyond the expected healing time of an injury and can involve structural changes and immune components. Additionally, chronic pain is often difficult to define and manage as it is individualized, often involving biological, emotional, and social aspects.
The “pain-on-a-chip” technology attempts to minimize this limitation by using biosensor systems to measure physiological response to a specific stimulus, which provides a clinician with an evidence-based alternative to verbal pain assessment. There is a microfluidic platform with specialized sensory cells that detect a change in calcium signalling in response to different chemicals that cause pain, such as ATP and capsaicin, which stimulate pain receptors.
The Pain-on-a-Chip device was constructed with AutoCAD and through photolithography on silicon wafers, providing an SU-8 photoresist pattern of 150 μm high patterns. A ratio of PDMS (polydimethylsiloxane) (10:1 base: curing) was used to create microfluidic chips following soft lithography, and curing was performed at 80 °C for 2 hours. After fabrication, the chips were used for the Pain-on-a-chip device by bonding to a glass coverslip using oxygen plasma treatment before adding rat tail collagen (40 μg/mL) to promote optimal cell attachment to each chip surface. Two designs were used: a single-channel chip (85 × 500 × 10,000 μm) and a multiplexed, three-channel chip (150 × 150 × 10,000 μm).
The 50 B11 neuronal cell line was seeded at a viable density of 1 × 10⁷ cells/mL concentration and differentiated for 48 hours in the presence of either NGF (nerve growth factor) or GDNF (glial cell line-derived neurotrophic factor) growth factors. Cells were adapted to the microfluidic chips by continuous supply of cell culture medium through a hydrated thread. Immunostaining of the cells was then done using specially designed anti-β III-tubulin primary and secondary antibodies, and then stained with Fura-2 AM (acetoxymethyl) to measure and follow intracellular Ca²⁺ concentrations during mechanochemical stimulation while immediately switching between 340 nm and 380 nm excitation wavelengths as well as 510 nm emission wavelength for the Ca²⁺ measurements.
To validate their device, the researchers conducted a series of tests that included immunostaining for neuronal markers and live imaging of calcium activity. The cells were exposed to known stimulants like ATP and capsaicin, which typically trigger distinct responses in pain-sensing neurons.
The implications for medical diagnostics are significant. At present, doctors rely on subjective scales to assess the presence of pain, which is a limited and laboured process. The consequent ability to objectively identify chronic pain types through a minimally invasive blood test could not only provide a validated diagnosis, but could delineate an individualised treatment plan, and objectively track outcome. The research team intends to enhance the platform with machine learning algorithms to improve the analysis of calcium signalling data.
A broader understanding of patterns of ion flux that correlate with the various pain syndromes faced by patients could allow AI to further enhance the biosensor’s predictive status and accuracy, building even more confidence in the device. In conclusion, it is apparent that the “pain-on-a-chip” biosensor represents a significant advance, and the first of its kind, in biomedical engineering, with applications for medical diagnostics. It combines microengineering with cellular biology and computational solutions into a single test. As the biosensor moves from the development stage into regulatory, it is hoped that the device will be a valuable tool in both clinical applications around pain management, but also preclinical drug discovery, and take aim at consideration for more individualised and more effective care for those who suffer from various chronic pain conditions.
References: Zhu D, Nilghaz A, Tong Z, et al. Pain-on-a-Chip: A microfluidic device for neuron differentiation and functional discrimination in animal models of chronic pain. Biosens Bioelectron. 2025;279:117401. doi:10.1016/j.bios.2024.117401
