In this Letter, the effects of Na+ and Mg2+ on the formation of triple helix DNA were examined by crosschecking of FRET signal from the different kinds of samples. Sodium abundant solution has not been thoroughly examined for the triple helix DNA experiment, although one can often face to the biological environment which has abundant Na+ ion. The result reconfirms that Mg2+ ion helps the oligonucleotides to form both double helix and triple helix DNA. It was conjectured and confirmed that Na+ plays a complex role, being configured by the polyelectrolyte effect and the effect of K+ on guanine-rich oligonucleotide. (C) 2010 Elsevier B.V. All rights reserved.