Deep eutectic solvents (DESs) have emerged as a promising class of environmentally friendly and sustainable solvents, offering an alternative to conventional organic solvents. Their distinctive features, including low cost, ease of preparation, tunable physicochemical properties, and high biodegradability, make them particularly attractive for applications in analytical chemistry. This chapter focuses on the expanding role of DESs in bioanalysis, especially in the context of sample preparation and analyte detection. Recent advancements and innovative methodologies employing DESs in the detection of various biological matrices, including blood, urine, hair, plasma, saliva, serum, sweat, and tears, are reviewed in detail. The chapter highlights the practical advantages of DES-based techniques over traditional solvent systems, such as enhanced extraction efficiency, improved selectivity, and reduced environmental impact. These developments position DESs as a powerful tool for future bioanalytical applications, bridging the gap between green chemistry and high-performance analysis.