Ethidium homodimer  (Synonyms: EthD-1)

Ethidium homodimer (EthD-1) is a high-affinity fluorescent nucleic acid dye commonly used to stain mammals, bacteria, yeast, and fungi. Ethidium homodimer binds to DNA or RNA, enhancing fluorescence more than 30 times. The Ethidium homodimer has a strong positive charge, so it cannot cross cell membranes and stain living cells; But the Ethidium homodimer can cross the disordered region of the dead cell membrane to reach the nucleus and embed the DNA double strand to produce red fluorescence. Therefore, Ethidium homodimer is a relatively sensitive nucleic acid stain that can accurately detect nucleic acids in solution or in decomposing cells. Ethidium homodimer binds DNA, Ex/Em=528/617 nm^[1].

Guide (The following is the experimental plan we recommend. This plan serves only as a reference guide. The specific operations should be adjusted according to your actual needs.)
1. Preparation of EthD-1 Working Solution 1.1 Preparation of Storage Solution Prepare 2 mM of EthD-1 using DMSO.
1.2 Preparation of Working Solution Dilute the stored solution with pre-warmed serum-free cell culture medium or PBS to prepare an EthD-1 working solution with a concentration of 0.1 - 10 μM.
Note: Please adjust the concentration of the EthD-1 working solution according to the actual situation, and prepare it as needed.

2. Cytological staining 2.1 Cell Preparation Suspended cells: Centrifuge the cells and then wash them twice with PBS, each time for 5 minutes.
Monolayer cells: Remove the culture medium and add trypsin to digest the cells. After centrifugation and discarding the supernatant, add PBS for two washes, each for 5 minutes.
Note: If no flow cytometry experiments are conducted, adherent cells can be subjected to slide staining.
2.2 Add 1 mL of EthD-1 working solution and incubate at room temperature for 15 to 60 minutes.
2.3 400 g, centrifuge at 4℃ for 3-4 minutes, then discard the supernatant.
2.4 Add PBS to wash the cells twice, for 5 minutes each time.
2.5 Resuspend the cells in 1 mL of serum-free medium or PBS, and then examine them under a fluorescence microscope or a flow cytometer.

Notes for Attention The EthD-1 storage solution is recommended to be aliquoted and stored under dark conditions at -20℃ or -80℃ to avoid repeated freezing and thawing. It can be stored for 1 month at -20℃ and for 6 months at -80℃.
2. Please adjust the concentration of EthD-1 working solution according to the actual situation.
3. This product is exclusively for professional scientific research purposes only. It shall not be used for clinical diagnosis or treatment, nor for food or medicine purposes.
4. For your safety and health, please wear a laboratory coat and put on disposable gloves when operating.

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

856.75

C₄₆H₅₀Cl₄N₈

61926-22-5

Solid

Red to reddish brown

NC1=CC=C2C(C(C3=CC=CC=C3)=[N+](CCCNCCNCCC[N+]4=C(C5=CC=CC=C5)C6=CC(N)=CC=C6C7=C4C=C(N)C=C7)C8=C2C=CC(N)=C8)=C1.Cl.Cl.[Cl-].[Cl-]

Room temperature in continental US; may vary elsewhere.

-20°C, sealed storage, away from moisture and light

*In solvent : -80°C, 6 months; -20°C, 1 month (sealed storage, away from moisture and light)

• [1]. Joshua R Edwards, et al. A novel method for the evaluation of proximal tubule epithelial cellular necrosis in the intact rat kidney using ethidium homodimer. BMC Physiol. 2007 Feb 23;7:1.  [Content Brief]