Sulfite (SO₃²⁻) is considered as a monitor of a wide range of physiological processes. However, cells and tissues are adversely affected when the body ingests high level of sulfite. Here, we designed and synthesized a “turn on” fluorescent probe ImiSft-1 with 2-cyano-N-methylacetamide as the specific recognition site of SO₃²⁻. This probe predominantly achieved high response intensity to SO₃²⁻ and desirable properties such as large Stokes shift (∼180 nm), fast response time (within 15 s), and high sensitivity (LOD = 0.12 μM). Importantly, the probe was highly selective for sulfite from other bio-species including biological thiols. Other functional properties included broad pH adaptability (5.0–10.0) and low cytotoxicity. Given these advantages and the fluorescence imaging in living MCF-7 cells, it was demonstrated that probe ImiSft-1 could monitor the changes of sulfite concentration in living cells.