Abstract
Tumor spheroids are one of the well-characterized 3D culture systems bearing close resemblance to the physiological tissue organization and complexity of avascular solid tumor stage with hypoxic core. They hold a wide-spread application in the field of pharmaceutical science and anti-cancer drug research. However, the difficulty in determining optimal technique for the generation of spheroids with uniform size and shape, evaluation of experimental outputs, or mass production often limits their usage in anti-cancer research and in high-throughput drug screening. In recent times, several studies have demonstrated various simple techniques for generating uniform-size 3D spheroids, including the hanging drop (HD), liquid overlay technique (LOT), and microfluidic approaches. Morphological alterations apart from biochemical assays, and staining techniques are suitably employed for the evaluation of experimental outcomes within 3D spheroid models. Morphological alterations in response to effective anti-cancer drug treatment in 3D tumor spheroids such as reduced spheroid size, loss of spheroid compactness and integrity or smooth surface, are highly reliable. These alterations can significantly reduce the need for biochemical assays and staining techniques, resulting in both time and cost savings. The present article specifically covers a variety of available procedures in spheroid generation. For practical applicability, we have supplemented our review study with the generation of glioblastoma U87 spheroids using HD and LOT methods. Additionally, we have also incorporated the outcome of U87 spheroid treatment with doxorubicin on spheroid morphology.