OD values are too low — close to background (CCK-8 Cell Viability Assay)

Question

Accepted Answer

Most OD values are below 0.1, the curve is essentially flat along the x-axis, and group differences are invisible. Common causes: 1) Seeding density too low 2) Culture period too short 3) Cell vitality is poor 4) CCK-8 reagent has expired/oxidized 5) Incubation time too short Solutions: 1) Increase seeding density appropriately 2) Extend culture time 3) Verify cell state and viability before plating 4) Use a fresh batch of CCK-8 reagent 5) Incubate at least 1 – 2 h before reading