Recently, there has been a growing interest in the use of enzymes as catalysts. Lipases thereby especially stand out while they are characterized by high activity in mild reaction conditions and a wide spectrum of reactions they catalyse. Therefore, it is crucial to produce the lipase of high catalytic activity which is commonly achieved by its purification. Enzyme purification is very often an expensive and slow procedure when conventional methods are used, so nowadays alternative, sustainable, and effective methods are being developed. The use of aqueous two-phase systems, that are recognized as green and cheap extraction agents, attracts a great interest in the purification of proteins and other molecules. An additional advantage is their significant flexibility, while aqueous two-phase systems can also be based on deep eutectic solvents which meet all the requirements and green chemistry principles due to their nontoxicity and biodegradability. For purification of sensitive biomolecules, such as proteins and enzymes, microsystems often stand out because they enable process intensification with a negligible disruption of molecule structures and carrying out a continuous process. In this paper, a method for continuous protein microextraction, namely lipase purification using a deep eutectic solvent based aqueous two-phase system was developed. After analysing the influence of deep eutectic solvent composition and salt concentration on extraction efficiency in the batch system, the system with betaine: urea deep eutectic solvent and K2HPO4 concentration of 0,7 g/mL proved to be optimal. The selected aqueous two-phase system was used for protein extraction in a microextractor where the influence of retention time, ultrasound, temperature, and channel diameter on the extraction process efficiency was examined. The optimal extraction efficiency achieved was 98,50 % for 30 s in a microsystem with a channel diameter of 1000 μm at 25 °C. The process was significantly intensified using a microextractor considering the extraction efficiency in a batch system which was 94,70 % for 30 min. The deep eutectic solvent reuse was also investigated, and it was successfully used in several extraction cycles in the batch system as well as in the microsystem. The developed microextraction method and the deep eutectic solvent capacity were further analysed using crude lipase produced by fermentation of Thermomyces lanuginosus on pumpkin oil pomace. Additionally, the microextraction process was successfully described by a 2D mathematical model.