经典的二维（2D）细胞培养作为药物或纳米颗粒测试系统，无法很好地复制体内条件。动物研究成本高昂，在伦理上存在争议，同时也限制了大规模测试的可能性。我们建立了一种三维（3D）组织切片空气液体界面（ALI）培养模型用于纳米颗粒测试。我们开发了一种优化程序，能够可靠地从肿瘤异种移植物中生成大批保留组织结构的组织切片。当用于基于化学改性聚乙烯亚胺（PEI）的纳米颗粒进行siRNA或DNA传递的分析时，相比于2D细胞培养，更清晰地观察到了纳米颗粒的转染效率和细胞毒性之间的差异。尽管纳米颗粒效果在细胞培养和组织切片模型之间总体上存在相关性，但组织切片模型还能够鉴定出特别合适的候选纳米颗粒，这些颗粒的功效在2D细胞培养中被低估，并且在之前的体内研究中已经得到证明。这种离体的3D组织切片ALI培养模型是一种强大的系统，它允许在完整的组织环境中有效评估生物纳米颗粒的功效和生物相容性。该系统成本相对较低、省时，遵循3R原则，可以鉴定关键的纳米颗粒性能和最佳的体内应用候选。©2022 Wiley-VCH GmbH出版的《生物技术杂志》

Classical two-dimensional (2D) cell culture as a drug or nanoparticle test system only poorly recapitulates in vivo conditions. Animal studies are costly, ethically controversial, and preclude large-scale testing.We established a three-dimensional (3D) tissue slice air-liquid interface (ALI) culture model for nanoparticle testing. We developed an optimized procedure for the reproducible generation of large sets of tissue slices from tumor xenografts that retain their tissue architecture. When used for the analysis of nanoparticles based on chemically modified polyethylenimines (PEIs) to deliver siRNA or DNA, differences in transfection efficacy and cytotoxicity between nanoparticles were observed more clearly than in 2D cell culture. While nanoparticle efficacies between cell culture and the tissue slice model overall correlated, the tissue slice model also identified particularly suitable candidates whose efficacy was underestimated in 2D cell culture and had already been shown in previous in vivo studies.The ex vivo 3D tissue slice ALI culture model is a powerful system that allows the effective evaluation of biological nanoparticle efficacy and biocompatibility in an intact tissue environment. It is comparably inexpensive, time-saving, and follows the 3R principle, while allowing the identification of critical nanoparticle properties and optimal candidates for in vivo applications.© 2022 The Authors. Biotechnology Journal published by Wiley-VCH GmbH.