A high-throughput workflow for assessing self-renewal using colony formation assays
Majd A Al-Hamaly1, Jessica S Blackburn2
1Pharmacology and Nutritional Sciences, University of Kentucky, Lexington, KY 40356, USA; Markey Cancer Center, University of Kentucky, Lexington, KY 40536, USA.
Abstract:
The colony formation assay (CFA) is a widely used method to assess the self-renewal capacity of cancer cells and evaluate how this property is affected by drug treatment. This protocol presents a streamlined, high-throughput CFA workflow optimized for T-cell acute lymphoblastic leukemia (T-ALL), making it suitable for large-scale drug screening projects. Colonies are grown in a methylcellulose-based 3D matrix and quantified using an automated analysis pipeline, allowing robust estimation of colony number and size. This cost-effective approach provides a scalable platform for identifying compounds that impair self-renewal, facilitating prioritization of hits for validation in more complex in vivo models.
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