What the best temperature and time for incubating cells using Hoechst #33342?

The optimal incubation time for Hoechst #33342 depends on the cells used and whether they are live or fixed.

• If the cells are fixed and permeabilized, the dye will stain the DNA quickly, within minutes. Temperature is not as important for fixed and permeabilized cells; room temperature or +4°C will produce good staining.
• For live cells, the incubation time will vary depending on the cell line. In general, 10-30 minutes at +37°C will provide good staining results.

Last updated: April 23, 2026