The abnormal changes of intracellular potassium ion (K+) concentration are usually associated with various diseases, therefore, real-time monitoring of intracellular K+ dynamic balance is crucial for disease research. Herein, we present a design strategy based on conjugated polymers (CPs) as potassium ion fluorescence probes. Six novel CP-based K+ sensors (P1, P2, P3, PP1, PP2 and PP3) were synthesized. These probes have different conjugated length backbones composing of stilbenzene, fluorene, thiophene, and/or benzothiadiazole units and different numbers of K+ selective ligands of triazacryptand (TAC), so as to study their spectral characteristics and response to K+. Among them, PP3 exhibits exceptional responsiveness and selectivity to K+. With a longer conjugated backbone, it shows a longer emission at 628 nm compared to other probes. Especially, the fluorescence intensities of PP3 showed a linear relationship with the concentrations of K+ in the range of 0–250 mM. PP3 has been successfully applied to detect the dynamic equilibrium of potassium ion concentration in living cells. This study proposes and validates the feasibility of the CP-based K+ fluorescence probe design strategy and expands the range of K+ probes, providing a new tool for understanding disease pathogenesis.