No colonies at all — total failure (Colony Formation Assay)

Question

Accepted Answer

After 7 – 14 days of culture, the entire dish shows no visible colonies after staining. Common causes: 1) Seeding density was too low (or no live cells survived) 2) Cell state / viability was very poor at seeding 3) Trypsin treatment was too long and damaged cells 4) Medium or serum is not suitable for clonogenic growth 5) Drug concentration in treatment group was too high (cytotoxic) Solutions: 1) Increase seeding density and run a pilot to find the right cell number 2) Use cells in log-phase with high viability (> 90 %) 3) Optimize trypsinization time (just enough to detach, no longer) 4) Switch to fresh medium / better serum batch 5) For treatment groups, lower drug concentration or reduce exposure time