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Longitudinal Bioluminescent Quantification of Three Dimensional Cell Growth
Michael K Wendt1, William P Schiemann1
1Case Comprehensive Cancer Center, Case Western Reserve University, Cleveland, USA.
Bio-Protocol
|July 26, 2016
Summary
This study introduces a novel assay using firefly luciferase to quantify three-dimensional (3D) breast cancer organoid growth. This high-throughput method enables longitudinal tracking, reducing costs and improving accuracy for 3D cell culture research.
Area of Science:
- Biomedical Engineering
- Cell Biology
- Cancer Research
Background:
- Three-dimensional (3D) cell culture models offer greater physiological relevance for mammary epithelial and breast cancer cells compared to traditional 2D systems.
- Existing methods for quantifying 3D organoid growth, such as [3H]thymidine, MTT assays, and area calculations, face significant experimental and technical limitations.
- Efficiently recovering cells from 3D matrices for analysis is time-consuming and can yield inaccurate results.
Purpose of the Study:
- To develop a novel, high-throughput assay for quantifying longitudinal cell outgrowth in 3D matrices.
- To overcome the limitations of current methods for assessing 3D mammary epithelial and breast cancer organoid growth.
- To provide a cost-effective and accurate method for tracking organoid development over time.
Main Methods:
- Stable expression of firefly luciferase in cells cultured within 3D matrices.
- Utilizing luminescence as a readout for quantifying cell outgrowth longitudinally.
- Implementing a high-throughput system for tracking single wells over time.
Main Results:
- The firefly luciferase assay enables accurate, longitudinal quantification of cell growth in 3D cultures.
- This method is high-throughput, allowing for efficient tracking of multiple samples simultaneously.
- The assay is cost-effective due to its ability to longitudinally monitor single wells.
Conclusions:
- The firefly luciferase assay offers a significant advancement for studying 3D mammary epithelial and breast cancer organoid growth.
- This technique facilitates the assessment of drug treatments and genetic manipulations on 3D organoid development.
- The assay's high-throughput nature and cost-effectiveness make it valuable for cancer research and drug discovery.
