Point‐of‐care testing (POCT) of clinical biomarkers is critical to health monitoring and timely treatment, yet biosensing assays capable of detecting biomarkers without the need for costly external equipment and reagents are limited. Blood‐based assays are, specifically, challenging as blood collection is invasive and follow‐upprocessing is required. Here, we report a versatile assay that employs hydrogel microneedles (HMNs) to extract interstitial fluid (ISF), in a minimally invasive manner integrated with graphene oxide‐nucleic acid (GO.NA)‐based fluorescence biosensor to sense the biomarkers of interest in situ. The HMN‐GO.NA assay is supplemented with a portable detector, enabling a complete POCT procedure. Our system could successfully measure four clinically important biomarkers (glucose, uric acid (UA), insulin, and serotonin) ex vivo, in addition, to accurately detecting glucose and UA in vivo.

Current methods for therapeutic drug monitoring (TDM) have a long turnaround time as they involve collecting patients' blood samples followed by transferring the samples to medical laboratories where sample processing and analysis are performed. To enable real‐time and minimally invasive TDM, a microneedle (MN) biosensor to monitor the levels of two important antibiotics, vancomycin (VAN) and gentamicin (GEN) is developed. The MN biosensor is composed of a hydrogel MN (HMN), and an aptamer‐functionalized flexible (Flex) electrode, named HMN‐Flex. The HMN extracts dermal interstitial fluid (ISF) and transfers it to the Flex electrode where sensing of the target antibiotics happens. The HMN‐Flex performance is validated ex vivo using skin models as well as in vivo in live rat animal models. Data is leveraged from the HMN‐Flex system to construct pharmacokinetic profiles for VAN and GEN and compare these profiles with conventional blood‐based measurements. Additionally, to track pH and monitor patient's response during antibiotic treatment, an HMN is developed that employs a colorimetric method to detect changes in the pH, named HMN‐pH assay, whose performance has been validated both in vitro and in vivo. Further, multiplexed antibiotic and pH detection is achieved by simultaneously employing the HMN‐pH and HMN‐Flex on live animals.

Diabetic ketoacidosis (DKA), a severe complication of type 1 diabetes (T1D), is triggered by production of large quantities of ketone bodies, requiring patients with T1D to constantly monitor their ketone levels. Here, a skin‐compatible hydrogel microneedle (HMN)‐continuous ketone monitoring (HMN‐CKM) device is reported. The sensing mechanism relies on the catechol–quinone chemistry inherent to the dopamine (DA) molecules that are covalently linked to the polymer structure of the HMN patch. The DA serves the dual‐purpose of acting as a redox mediator for measuring the byproduct of oxidation of 3‐beta‐hydroxybutyrate (β‐HB), the primary ketone bodies; while, also facilitating the formation of a crosslinked HMN patch. A universal approach involving pre‐oxidation and detection of the generated catechol compounds is introduced to correlate the sensor response to the β‐HB concentrations. It is further shown that real‐time tracking of a decrease in ketone levels of T1D rat model is possible using the HMN‐CKM device, in conjunction with a data‐driven machine learning model that considers potential time delays.