Vital Staining of Chromosomes with Hoechst 33258
Materials
1. Hoechst 33258, 10 mg/ml stock in DMSO.
2. Culture medium.
3. 15 ml sterile conical tube.
Procedure
1. In a 15 ml conical tube, add 1 volume Hoechst 33258 stock to 1000 volumes of medium. Need 3 ml for each coverlisp dish. Mix thoroughly.
2. Remove all medium from the culture dish, replace with the dye-containing medium.
3. Incubate for 30 min.
4. Remove dye-containing medium. Rinse dish twice with regular medium.
5. Add regular medium to the dish. Fluorescence can be observed with Zeiss 02 UV filter set and quartz-halogen lamp. Mercury arc lamps yield a much higher intensity but could cause damange to live cells.
Note: the same protocol can be used with ethidium bromide. Replace Hoechst stock solution with 10 mg/ml aqueous stock solution of ethidium bromide.
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