Cell of the Month: Cells in a 3D Spheroid

Jun 9, 2020 | Disease Models

Spheroids have been used in cell culture for decades. In the 1980s, different types of human cancer cells –normally grown as monolayers or suspension cultures–were tested for their innate abilities to form and grow as spheroids in vitro (1). 16 out of 27 tested tumor cell lines successfully formed spheroids (1). In the tumor cells study, scientists also observed that all large spheroids had necrotic centers but their shapes varied. Another study used V79 hamster cells (V79 379A), a human small cell carcinoma of the lung (ME/MAR) and 2 xenographed human melanomas (HX117 and HX118) in spheroids cultures in the 1980s (2). In the study, the effects on spheroid growth due to radiation treatments were measured and evaluated (2).

Widely acknowledged is the unique ability of spheroids to mimic natural cell responses and interactions. Cells in 3D are more representative of their native conditions than the traditional 2D monolayer culturing conditions (e.g., cell-to-cell interactions, drug-induced responses, and cells-to-environment responses). Furthermore, the multicellular arrangement allows different cell types to interact with each other within each spheroid. Past studies examined growth rates, hypoxic conditions, and other survival conditions for the spheroids (1-2). Currently, many cell culture reagents and instruments are available to enable spheroid cultures, making the technique increasingly accessible, flexible and approachable for scientists in a variety of research fields. (Unlock your creative minds!)

In 2019, drug induced activities were conducted using 3D cell cultures (3). Modeling rare types of pancreatic tumors, multiple 3D cell culture techniques were compared: Method A: hanging drop plates; Method B: 24-well plates with cell repellent surface; and, Method C: ultra-low attachment 96-well plates. The results showed that Method C (ultra-low attachment wells and plates) was the most effective approach using the following evaluation criteria: cell seeding, morphology, culture medium maintenance, quality of spheroids, reproducibility and overall time required for the experiments (3).

At Tempo Bioscience, we recommend the ultra-low attachment wells and plates approach for making 3D spheroids. Our favorite examples are : 1) neural-glial spheroids using Tempo-iCort, Tempo-iAstro, Tempo-iOligo, and Tempo-iMG; and 2) kidney podocytes spheroids using Tempo-iKidneyPod. (Read more about human podocytes here).

Email Tempo Support for 3D spheroid cell culture protocols!


1) Carlsson J., et al Formation and Growth of Multicellular Spheroids of Human Origin. Int J Cancer 1983 May 15;31(5):523-33. doi: 10.1002/ijc.2910310502.

2) West, CM., Sandhu, RR., and Stratford, IJ. The radiation response of V79 and human tumour multicellular spheroids–cell survival and growth delay studies. Br J Cancer. 1984 Aug; 50(2): 143–151. doi: 10.1038/bjc.1984.156.

3) Bresciani, G., et al. Evaluation of Spheroid 3D Culture Methods to Study a Pancreatic Neuroendocrine Neoplasm Cell Line. Front Endocrinol (Lausanne). 2019; 10: 682. doi: 10.3389/fendo.2019.00682.