Tumorsphere Formation Assay

Overview

The tumorsphere formation assay measures the ability of cancer cells to grow as non-adherent spherical colonies under non-adherent, serum-free conditions. Tumorsphere-forming capacity is a functional surrogate for cancer stem-like cell (CSC) properties, as only cells with self-renewal capacity can survive and proliferate without anchorage. The number and size of spheres formed reflects stemness potential.

Used by

• Used in GBC cell lines (GBC-SD, NOZ) to assess stemness; conditioned medium from stiff-matrix-activated gallbladder fibroblasts increased tumorsphere formation, an effect abrogated by SEMA7A knockdown in fibroblasts and rescued by recombinant human SEMA7A; downstream p300 inhibition (C646 or shRNA) also blocked sphere formation PMID:24997986

Notes

• Typically performed in ultra-low attachment plates with serum-free medium supplemented with EGF and bFGF.
• Sphere efficiency (number of spheres / cells seeded × 100%) is the standard readout.
• Commonly used alongside invasion assays to jointly assess EMT and stemness phenotypes.

Sources

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