The growing prevalence of chronic digestive disorders, such as inflammatory bowel disease, underscores the urgent need for innovative solutions that enable longitudinal monitoring of disease progression and treatment efficacy. Addressing this challenge, we present a novel microneedle-based sensor designed for rapid, point-of-care assessment of intestinal barrier integrity. Through transient application to the skin, the device samples intestinal fatty acid binding protein (IFABP) from systemic circulation, offering a minimally invasive alternative to conventional diagnostics. We demonstrate a versatile, affinity-based electrochemical sensing mechanism integrated into low-cost and clean room-free microneedles. The resulting device is validated in a biomimetic skin-like hydrogel in which it achieves good linearity, a limit of detection of 1.5 ng mL−1 and highly specific response in a short assay format of one hour including the sampling phase. Furthermore, we validate the sensor's biocompatibility, penetration efficiency, and sensing capability in ex vivo human skin, establishing a critical foundation for future clinical applications. This breakthrough technology holds significant promise for transforming the management of gastrointestinal diseases through frequent, patient-friendly monitoring.