**1. Introduction**

Nucleotide adenosine triphosphate (ATP) is one of the critical biological anions in all living organisms. It serves as a universal energy source in various cellular events and plays significant roles in many biological processes including biosynthesis, DNA replication, cell signaling, and so on [1–5]. The intracellular ATP levels have been demonstrated to be closely relevant to pathogenesis of many diseases including Parkinson's disease, ischemia, and hypoglycemia, etc. [5–8]. Therefore, sensitive and selective detection of ATP is highly desired for biochemical research and clinical diagnosis.

In recent years, fluorescent probes are considered as powerful tools for the visible detection and identification of biological substances owing to the good selectivity, high sensitivity, real-time analysis capability, and high temporal and spatial resolution [9–14]. Thanks to the strong light-harvesting and signal-amplification properties of conjugated polymers (CPs), water-soluble CPs-based probes have been demonstrated to be efficient probes for detection of diverse bio-related species such as nucleotides, DNA, proteins, protease, etc. [15–17]. Among various CPs, polythiophene (PT) derivatives have received extensive attention because of their sensitive chain conformation transition responsive to external stimuli [7,8,18–21], providing unique advantages over other CPs on keeping the balance between simplicity (colorimetric mode) and sensitivity (fluorescence mode). In previous reports including ours, several PTs derivatives have been designed for the fluorescent detection of ATP with moderate selectivity and sensitivity [19,22–25]. These reported PT-based ATP probes contain generally cationic groups on their side chains, for example, quaternary ammonium [18,19,25,26], imidazolium [22], or phosphonium [23], to offer essential water solubility and recognition site for ATP. Thus, in most cases, electrostatic interaction is the primary driving force for the binding of polymer probes with ATP, resulting in these assays being sensitive to ionic strength. This will prevent practical application of these probes in more complex environments like cells and even living body. On the other hand, the selectivity of the reported ATP probe was not satisfying since the electrostatic attraction was nonspecific. To address these challenges, herein, we propose a multisite-binding coupled with analyte-induced aggregation strategy to develop PT-based probes for highly selective ATP detection and imaging.

It is well known that boronic acids can bind vicinal diols with high a ffinities via reversible boronate formation in weak-base aqueous solution, which has been widely used to develop synthetic receptors for saccharides detection [6,26–29]. In the present contribution, both boronic acid groups and quaternary ammonium groups were introduced to the side chain of polythiophene as binding sites of ribose and phosphate of ATP, respectively, to produce a new ATP probe (**L**) with strong and selective affinity for ATP (Scheme 1a). **L** and ATP come together easily by the electrostatic attraction, at the same time, the neighboring boronic acid group on the side chains of **L** further anchors ATP via formation of pentacyclic borate to restrict the rotation of **L** backbone and thus, promote polymer aggregation through the interchain π–π stacking interaction (Scheme 1b). This process will lead to the significant fluorescence quenching of **L**. Based on the sensing strategy, **L** can distinguish ATP from its analogues ADP and AMP as well as various inorganic phosphates, etc. Taking advantage of high selectivity of **L** to ATP, the activity assay of ATP-related enzyme in bu ffer solution and the fluorescence imaging of ATP in living cells were successfully achieved.

**Scheme 1.** (**a**) The possible binding pattern of **L** with ATP; (**b**) Conformation transition and the formation of aggregates of **L** induced by ATP.
