Bioelectronics is an evolving field focused on creating efficient interfaces between electronic systems and biological environments, where electrodes are crucial for detecting, recording, and stimulating signals. While traditional fabrication techniques offer high precision, they are costly and lack scalability. Vacuum soft lithography offers a scalable, time-efficient alternative for manufacturing microstructured electrodes. When combined with poly(3,4-ethylenedioxythiophene) poly(styrene sulfonate) (PEDOT:PSS), a conducting polymer known for its biocompatibility and stability, this approach presents a promising solution for advancing bioelectronic applications. This study demonstrates the successful application of vacuum soft lithography for fabricating PEDOT:PSS-based microstructured electrodes. These PEDOT:PSS-based microstructured electrodes are fabricated and characterized based on their morphology, mechanics, and electrochemical properties. The PEDOT:PSS mixture containing 2 % (3-glycidyloxypropyl)trimethoxysilane (GOPS) demonstrates optimal properties, achieving an excellent balance of reproducibility, conductivity, and mechanical integrity, with in vitro cytocompatibility confirmed through testing with the MDA-MB-231 breast cancer cell line. This approach offers significant advantages, including reduced costs and production times, making it a viable alternative to conventional methods. Future research will explore its use with other conducting materials for broader bioelectronic applications.