Preparation of 1 mM EDTA solution from stock solution of EDTA (0.5M, pH 8.0)


  • EDTA (Ethylenediaminetetraacetic acid) is a chelating agent. Due to its ability to sequester metal ions such as Mn2+, Ca2+, Mg2+ and Fe3+, EDTA is widely used in number of cell and molecular biology experiments.
  • Metal ions are necessary for the activity of many enzymes (e.g., DNases and DNA modifying enzymes) as well as interactions of biomolecules (e.g., receptor-ligand interaction). Sequestration of metal ions disrupts metal ion-dependent interactions (e.g., cell-cell and cell substratum interaction) as well as inhibits the action of metal ion dependent enzymes.
  • Here we describe steps to prepare 1 mM EDTA solution from stock EDTA solution.


  • Reagents
    • 0.5 M EDTA solution, pH 8.0 at 25°C
    • Deionized / Milli-Q water
  • Equipment and disposables
    • Measuring cylinder


1 mM  EDTA, pH 8.0 at 25°C


Preparation of 100 ml of 1 mM EDTA solution (pH 8.0) from 0.5 M EDTA solution (pH 8.0)


Step 1: Calculate amount of 0.5M EDTA required for the preparation of 100 ml of 1 mM EDTA.

You need to add 0.2 ml of 0.5 M EDTA in 99.8 ml water to achieve 100 ml of 1 mm EDTA solution. (See calculation)

Step 2: Take ≈50 ml water in a measuring cylinder and add 0.2 ml of 0.5M EDTA in it.

Step 3: Adjust the final volume 100 ml with deionized/milli Q water. Mix it.

Tips: To mix solution, you can cover the measuring cylinder with parafilm and mix the solution by inverting the measuring cylinder 3 – 4 times. Take care no solution leaks out during this process. Alternatively, you can transfer solution to storage bottle and mix solution by swirling the bottle or using magnetic stirrer.


Transfer solution to a storage bottle. You can store the solution at room temperature. Solution is stable at room temperature for long time.


Follow the table to prepare EDTA solution of specific concentration and volume from 0.5 M EDTA solution
Conc. / Volume 100 ml 250 ml 500 ml 1000 ml
1 mM 0.2 ml 0.5 ml 1.0 ml 2.0 ml
5 mM 1.0 ml 2.5 ml 5.0 ml 10.0 ml
10 mM 2.0 ml 5.0 ml 10.0 ml 20.0 ml
25 mM 5.0 ml 12.5 ml 25.0 ml 50.0 ml