Infertility remains a significant global health challenge, and while conventional assisted reproductive technologies (ARTs) are effective, they are often limited by mechanical stress and the bypassing of natural physiological selection barriers. This review examines the application of microfluidics to prepare gametes, offering a solution to these limitations. By leveraging biological mechanisms such as rheotaxis, thigmotaxis, chemotaxis, and thermotaxis, microfluidic systems enable the non-invasive, selective isolation of high-quality sperm. Furthermore, this technology mimics the in vivo environment more accurately, optimizing sensitive processes like enzyme-free oocyte denudation and in vitro maturation (IVM). This approach minimizes cytoplasmic damage and preserves genetic integrity. Finally, dynamic embryo culture on microfluidic platforms may provide a continuous supply of nutrients and efficient removal of metabolic waste, thereby enhancing fertilization rates and blastocyst formation compared to static methods.