What is the best concentration to use with Hoechst?

Hoechst can be used at concentrations of 1-10 ug/mL in phosphate buffered saline (PBS) or any other detergent-free buffer at room temperature. The best concentration for staining live cells with Hoechst is 1 ug/mL. Cytotoxicity is minimum at this concentration, ensuring a higher viability of stained live cells. Over-staining with Hoechst is very rare because the dye has limited binding sites.