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Cell Viability Assays in Three-Dimensional Hydrogels: A Comparative Study of Accuracy
Anthony J Dominijanni1, Mahesh Devarasetty1, Steven D Forsythe1
1Wake Forest Institute for Regenerative Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA.
Tissue Engineering. Part C, Methods
|June 4, 2021
Summary
Researchers evaluated common cell viability assays for accuracy in 3D cell cultures, finding inaccuracies. Microscopic imaging is recommended for validating results from these 3D cell culture models.
Area of Science:
- Biotechnology
- Tissue Engineering
- Cell Biology
Background:
- Three-dimensional (3D) cell culture models, like organoids, offer physiological accuracy for drug development and precision medicine.
- Advances in tissue engineering and biofabrication enable complex 3D systems, but quantitative cell viability analysis remains a challenge.
- Existing cell viability assays, developed for 2D cultures, are increasingly applied to 3D constructs, raising questions about their accuracy.
Purpose of the Study:
- To compare the accuracy of commercially available cell viability assays for use in 3D hydrogel-based cell culture systems.
- To identify potential inaccuracies when using 2D-designed viability assays in diverse 3D hydrogel formulations (organic, synthetic, hybrid).
- To provide guidance for researchers on assay selection and validation in 3D cell culture experiments.
Main Methods:
- Commercially available viability assays (Promega's CellTiter-Glo®, CellTiter-Glo 3D, CellTiter 96® MTS; Thermo Fisher's PrestoBlue™) were tested on 3D hydrogel constructs.
- Assay performance was evaluated across different hydrogel compositions.
- Results were compared against direct cell viability measurements typically used in 2D cultures.
Main Results:
- Significant cellular health output inaccuracies were observed for the tested assays across various hydrogel formulations.
- The study highlights that assays designed for 2D cultures may not accurately reflect cell viability in 3D environments.
- Variability in results necessitates careful consideration when interpreting cell viability data from 3D cultures.
Conclusions:
- Researchers must be aware of potential errors when applying standard 2D cell viability assays to 3D cell culture systems.
- Microscopic imaging should be employed in conjunction with viability assays for accurate validation of cellular health in 3D constructs.
- This study informs the scientific community about the limitations of current viability assays in 3D models, promoting more reliable experimental outcomes.
