Wearable health monitoring devices are gaining significant traction, with a growing interest in real-time monitoring of metabolic status via easily accessible biological fluids like sweat. Sweat is particularly appealing due to its non-invasive accessibility and richness in physiological and metabolic biomarkers that correlate well with blood plasma levels. These biomarkers, including sodium (hydration status), lactate (muscle fatigue), chloride (cystic fibrosis diagnosis), cortisol (stress), glucose (energy levels, diabetes), and ammonium (metabolic conditions), offer valuable insights into human health and performance. Real-time, non-invasive biosensors capable of simultaneously detecting multiple biomarkers are crucial for accurate diagnosis and personalized health management. While various sweat sensor formats exist (tattoos, sweatbands), textile-based approaches offer superior versatility and breathability, enabling seamless integration into existing clothing. However, existing textile-based electrochemical sensors often suffer from non-uniformity and variability due to ink wicking on uneven fabric surfaces. This research addresses these challenges by utilizing a bottom-up approach with individual functionalized threads, offering adaptability and ease of integration into any garment. This manuscript details a fully integrated, wearable, thread-based multiplexed platform for real-time detection of sodium, ammonium, pH, and lactate directly from sweat, with wireless electronic readout. The chosen biomarkers—sodium, lactate, ammonium, and pH—provide a real-time physiological measure of athletic performance and physical activity. Sodium indicates hydration, lactate reflects muscle fatigue (also monitored by pH), and ammonium reveals anaerobic metabolism. The platform utilizes polyester (PE) and stainless steel (SS) threads coated with conductive inks as sensing electrodes. Potentiometric detection of electrolytes and pH is achieved through ion-selective membrane deposition and pH-sensitive polyaniline coating, while amperometric lactate detection employs a lactate oxidase-based enzymatic sensing scheme. The thread sensors are integrated onto a patch connected to a miniaturized circuit module with a potentiostat, microprocessor, and wireless circuitry for smartphone readout.

The literature review highlights the growing interest in minimally invasive wearable health monitoring devices and the advantages of using sweat as a readily accessible and informative biological fluid. Existing sweat sensors have been developed in various formats, including tattoos, sweatbands, and wristbands, utilizing different substrates such as tattoo paper, cellulosic paper, and flexible polymers. However, challenges remain in integrating electronics with flexible sensors and achieving real-time, continuous sweat analyte measurement. Textile-based approaches have emerged as a promising solution due to their versatility and breathability, allowing for integration into existing clothing. While previous research explored textile-based sensors for perspiration rate, pH, and lactate, these often relied on screen-printing or dip-coating, resulting in inconsistencies. The authors’ previous work on thread-based microfluidic networks and sensors provided the foundation for this study, demonstrating the potential of functionalized threads for various applications, including wound pH monitoring and on-demand drug delivery. The inherent properties of threads (flexibility, tunability, strength, inertness) make them ideal substrates for wearable sensor platforms, surpassing the limitations of conventional on-fabric approaches.

The methodology section details the fabrication and characterization of the thread-based sensors and the integrated wireless readout system. Conductive threads were created by coating PE and SS threads with conductive inks (carbon and Ag/AgCl). For potentiometric sensing of electrolytes and pH, ion-selective polymeric membranes and polyaniline coatings were applied. Amperometric lactate sensing was achieved using enzyme-coated threads (lactate oxidase on Prussian blue-coated threads). The electrical conductivity of the coated threads was characterized, showing optimal resistance after three coatings. Scanning electron microscopy (SEM) confirmed the uniform coating of the sensing layers. Individual sensor performance was evaluated using standardized analyte solutions, demonstrating near-Nerstian behavior for the potentiometric sensors and high sensitivity for the amperometric lactate sensor. The response time (5-30 s) was deemed suitable for real-time monitoring. Hysteresis tests showed reproducible responses without rinsing, validating the sensors’ suitability for continuous sweat measurement. Selectivity testing confirmed minimal interference from other electrolytes. Long-term drift analysis showed minimal response variation over three hours. The wireless readout electronics were designed to minimize size, weight, area, and power consumption (SWAP). A low-power integrated circuit with wireless configuration capabilities was employed, enabling simultaneous potentiometric and amperometric measurements. A graphical user interface (GUI) facilitated system programmability, data acquisition, and visualization. In vivo testing involved on-body trials with human volunteers performing exercise (stationary cycling and treadmill running). Initial trials used a commercial potentiostat, while subsequent trials employed the custom wireless readout electronics. Data were collected during various exercise intensities, capturing real-time changes in sweat biomarkers. Maximal oxygen uptake (VO2max) tests were conducted with some participants to correlate sweat biomarker levels with physical fitness. The data was then processed and converted into concentrations using the earlier calibration plots. A detailed description of materials and instrumentation used is also provided, including information on specific chemicals, inks, and equipment used in the fabrication and characterization processes.

The key findings demonstrate the successful fabrication and validation of a fully integrated, wearable, thread-based multiplexed sweat sensor patch. The potentiometric sensors exhibited near-Nerstian behavior with high sensitivities for sodium (52.8 mV/decade), ammonium (60.6 mV/decade), and pH (62.3 mV/decade). The amperometric lactate sensor showed a sensitivity of 900 nA/mM. Response times were within the range of 5-30 s. In vitro and in vivo experiments confirmed the sensors' ability to detect real-time changes in sweat analytes during exercise. The custom wireless readout electronics module successfully collected multiplexed data from the sensors. On-body trials during stationary cycling and treadmill running demonstrated the system’s capability to track changes in sodium, ammonium, pH, and lactate concentrations in response to exercise intensity. The data obtained during the VO2max tests showed correlations between VO2 and sweat analyte levels, indicating the potential for real-time monitoring of physical fitness and fatigue. Specifically, for Subject 6, higher VO2 was associated with higher sodium (r=0.847, p < 0.001) and lower ammonium (r = -0.785, p=0.001). For Subject 7, higher VO2 was associated with higher pH (r=0.773, p=0.003) and lower ammonium (r=-0.812, p=0.001), but not sodium (p=0.064). These correlations support the utility of this sweat sensing platform for assessing aspects of physical fitness and endurance.

The findings address the research question by demonstrating the feasibility of a wearable, multiplexed sweat sensor patch for real-time monitoring of key physiological biomarkers. The sensors' performance is comparable to other established sensor technologies, and the system successfully captures the dynamic changes in sweat composition during exercise, which is useful for understanding physical performance and fatigue. The correlation between VO2max and sweat analytes, although based on a limited sample size, indicates the platform's potential for assessing physical fitness. The platform’s potential extends beyond athletic performance monitoring; the real-time multiplexed detection of sweat biomarkers opens opportunities for clinical diagnostics and personalized health management. Future studies should focus on expanding the range of detectable biomarkers and improving the system's robustness to various environmental conditions. Advanced signal processing techniques can be employed to mitigate motion artifacts during vigorous exercise. Larger-scale clinical studies are needed to validate the system's accuracy and reliability for diagnostic purposes.

This study successfully demonstrates a cost-effective, flexible, and wearable multiplexed platform for real-time sweat monitoring. The thread-based sensor patch, integrated with wireless electronics, allows for simultaneous and continuous detection of key biomarkers related to hydration, muscle fatigue, and anaerobic metabolism. The initial findings show promising correlations between these biomarkers and physical fitness, paving the way for applications in athletic performance monitoring and potentially clinical diagnostics. Future work will focus on larger-scale studies, improved signal processing, miniaturization of the electronics, and expansion of the detectable biomarker panel.

The study has limitations in terms of the relatively small sample size of the in vivo trials. This limits the statistical power of correlations observed between VO2max and sweat analyte levels. Further, the integration of the custom electronics module with the patch could be improved to reduce motion artifacts completely and improve the signal quality during vigorous exercise. While the sensor showed good stability over 3 hours, longer-term studies are needed to evaluate the sensors' long-term stability and durability. The study primarily focused on healthy individuals and additional studies are needed to determine the platform’s applicability to diverse populations and clinical conditions.