Because of the many advantages they offer (strength, low cost, durability, lightweight, resistance, etc.), plastics are integral part of our daily life with a production constantly rising. However, their waste management is still inadequate, resulting in their release and accumulation in the environment, representing a main source of pollution. Their degradation results in debris of variable size including microplastics (0.1 μm-5 mm) and even nanoplastics (<0.1 μm), whose potential impact on ecosystems and human health have raised concerns. The potential adverse effects they may cause have been evaluated using both in vitro and in vivo models. However, due to some specific characteristics of micro- and nanoplastics, there are challenging questions about whether conventional in vitro tests are appropriate for evaluating their toxicity. For example, low-density plastics float on the surface of the culture medium and cannot come into contact with cells adhering to the bottom of the culture plates, which prevents proper evaluation of potential adverse effects and leads to misinterpretation of toxicological assays. In this review, we discuss the main issues related to the evaluation of micro- and nanoplastics toxicity using conventional in vitro assays. A literature survey has allowed to propose some solutions to circumvent these issues including the use of mathematical models to accurately determine the dose of particles delivered to cells, advanced 3D models (organoids), inverted cell culture models, cell cultures at the air-liquid interface or under dynamic conditions. Finally, we propose some perspectives and recommendations for further research on the in vitro evaluation of micro- and nanoplastics toxicity, underlining the importance of using standardized protocols for comparison purposes and samples and experimental conditions more representative of real-life exposure.