title: Reproducibility of Uniform Spheroid Formation in 384-Well Plates: The Effect of Medium Evaporation.
abstract: Spheroid cultures of cancer cells reproduce the spatial dimension-induced in vivo tumor traits more effectively than the conventional two-dimensional cell cultures. With growing interest in spheroids for high-throughput screening (HTS) assays, there is an increasing demand for cost-effective miniaturization of reproducible spheroids in microtiter plates (MPs). However, well-to-well variability in spheroid size, shape, and growth is a frequently encountered problem with almost every culture method that has prevented the transfer of spheroids to the HTS platform. This variability partly arises due to increased susceptibility of MPs to edge effects and evaporation-induced changes in the growth of spheroids. In this study, we examined the effect of evaporation on the reproducibility of spheroids of tumor and nontumor cell lines in 384-well plates, and show that culture conditions that prevent evaporation-induced medium loss result in the formation of uniform spheroids across the plate. Additionally, we also present a few technical improvements to increase the scalability of the liquid-overlay spheroid culturing technique in MPs, together with a simple software routine for the quantification of spheroid size. We believe that these cost-effective improvements will aid in further improvement of spheroid cultures for HTS drug discovery.
authors.affiliation: Institute of Molecular and Translational Medicine, Faculty of Medicine and Dentistry, Palacky University, Olomouc, Czech Republic.
authors.forename: Viswanath
authors.initials: V
authors.lastname: Das
authors.affiliation: Institute of Molecular and Translational Medicine, Faculty of Medicine and Dentistry, Palacky University, Olomouc, Czech Republic.
authors.forename: Tomáš
authors.initials: T
authors.lastname: Fürst
authors.affiliation: Institute of Molecular and Translational Medicine, Faculty of Medicine and Dentistry, Palacky University, Olomouc, Czech Republic.
authors.forename: Soňa
authors.initials: S
authors.lastname: Gurská
authors.affiliation: Institute of Molecular and Translational Medicine, Faculty of Medicine and Dentistry, Palacky University, Olomouc, Czech Republic.
authors.forename: Petr
authors.initials: P
authors.lastname: Džubák
authors.affiliation: Institute of Molecular and Translational Medicine, Faculty of Medicine and Dentistry, Palacky University, Olomouc, Czech Republic marian.hajduch@upol.cz.
authors.forename: Marián
authors.initials: M
authors.lastname: Hajdúch
mesh_terms: D000970:Antineoplastic Agents / Q000302:isolation & purification* / Q000627:therapeutic use; D018929:Cell Culture Techniques / Q000379:methods*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D055808:Drug Discovery / Q000379:methods*; D004354:Drug Screening Assays, Antitumor / Q000379:methods; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D018874:Spheroids, Cellular / Q000187:drug effects
chemical_list: D000970:Antineoplastic Agents
keywords: edge effect; evaporation; liquid-overlay culture; reproducibility; spheroids