Microfluidics has emerged as a valuable tool for the high-resolution patterning of biological probes on solid supports. Yet, its widespread adoption as a universal biological immobilization tool is still limited by several technical challenges, particularly for the patterning of isolated spots using three-dimensional (3D) channel networks. A key limitation arises from the difficulties to adapt the techniques and materials typically used in prototyping to low-cost mass-production. In this paper, we present the fabrication of thin thermoplastic elastomer membranes with microscopic through-holes using a hotembossing process that is compatible with high-throughput manufacturing. The membranes provide the basis for the fabrication of highly integrated 3D microfluidic devices with a footprint of only 1 Â 1 cm 2 . When placed on a solid support, the device allows for the immobilization of up to 96 different probes in the form of a 10 Â 10 array comprising isolated spots of 50 Â 50 mm 2 . The design of the channel network is optimized using 3D simulations based on the Lattice-Boltzmann method to promote capillary action as the sole force distributing the liquid in the device. Finally, we demonstrate the patterning of DNA and protein arrays on hard thermoplastic substrates yielding spots of excellent definition that prove to be highly specific in subsequent hybridization experiments.