LAUSR

Catalytic DNA‐based fluorescent sensors have enabled cellular imaging of metal ions such as Mg2+. However, natural DNA is prone to nuclease‐mediated degradation. Here, we report the in vitro selection of threose nucleic acid enzymes (TNAzymes) with RNA endonuclease activities. One such TNAzyme, T17–22, catalyzes a site‐specific RNA cleavage reaction with a kcat of 0.017 min−1 and KM of 675 nM. A fluorescent sensor based on T17–22 responds to an increasing concentration of Mg2+ with a limit of detection at 0.35 mM. This TNAzyme‐based sensor also allows cellular imaging of Mg2+. This work presents the first proof‐of‐concept demonstration of using a TNA catalyst in cellular metal ion imaging.