Until recently, chemiluminescence cell images could only be obtained using luciferase-activated probes. Moreover, chemiluminescence microscopy cell-imaging has not been demonstrated for natively expressed enzymes like cathepsin B. Herein, we describe the design, synthesis, and evaluation of the first chemiluminescence probe for the detection and imaging of cathepsin B. The probe activation mechanism relies on the release of a dioxetane intermediate, which undergoes chemiexcitation to emit green light with high efficiency under physiological conditions. Using the probe, we obtained clear images of cancerous leukemia and colon cells. This is the first demonstration of chemiluminescence cell images obtained by a probe for a natively expressed endogenous enzyme. We anticipate that the concept presented in this study will be broadly used to develop analogous probes for other important proteases relevant to biomolecular processes.

Detection of Salmonella and L. monocytogenes in food samples by current diagnostic methods requires relatively long time to results (2-6 days). Furthermore,t he ability to perform environmental monitoring at the factory site for these pathogens is limited due to the need for laboratory facilities. Herein, we report new chemiluminescence probes for the ultrasensitive direct detection of viable pathogenic bacteria. The probes are composed of ab right phenoxy-dioxetane luminophore masked by triggering group,whichisactivated by aspecific bacterial enzyme,and could detect their corresponding bacteria with an LOD value of about 600-fold lower than that of fluorescent probes.M oreover,w ew ere able to detect am inimum of 10 Salmonella cells within 6hincubation. The assayallows for bacterial enrichment and detection in one test tube without further sample preparation. We anticipate that this design strategy will be used to prepare analogous chemiluminescence probes for other enzymes relevant to specific bacteria detection and point-of-care diagnostics.

Low‐molecular‐weight self‐assembled peptides may serve as promising hydrogelators for drug delivery applications by changing their structural network in response to external stimuli. Herein, inspired by the well‐studied low‐molecular‐weight peptide hydrogelator, fluorenyl‐methoxycarbonyl‐diphenylalanine (Fmoc‐FF), a novel peptide is designed and synthesized to include an ultraviolet (UV)‐sensitive phototrigger. Similar to Fmoc‐FF, 6‐nitroveratryloxycarbonyl‐diphenylalanine (Nvoc‐FF) self‐assembles to form a 3D, self‐supporting, nanofibrous hydrogel. The Nvoc‐FF hydrogel exhibits good mechanical properties with a storage modulus of 40 kPa. UV irradiation of the Nvoc‐FF hydrogel encapsulating insulin‐fluorescein isothiocyanate (insulin‐FITC) results in the cleavage of Nvoc‐FF peptide to produce unmasked FF, thereby facilitating the degradation of the hydrogel and the release of insulin‐FITC. This release is in linear correlation to the irradiation time. In the present study, a first insight into this rigid, fibrous, light‐responsive hydrogel is provided, allowing the fabrication of a novel drug delivery system for controlled release of large molecules.

A recent methodology, developed by our group, has enabled a dramatic improvement in the emissive nature of the excited species, formed during the chemiexcitation of dioxetanes under physiological conditions. This approach has resulted in the discovery of distinct phenoxy-dioxetane luminophores that produce a chemiluminescence signal via a direct-mode of emission. Here, we show a significant pK effect of our new phenoxy-dioxetanes on their chemiexcitation and on their ability to serve as chemiluminescent turn-ON probes for biological applications. Using an appropriate phenoxy-dioxetane probe with a direct-mode of emission, we were able to image β-galactosidase activity, in cancer cells and in tumor-bearing mice. To the best of our knowledge, this is the first example to demonstrate in vitro and in vivo endogenous enzymatic chemiluminescence images obtained by a single-component phenoxy-dioxetane probe. We anticipate that our strategy, for the design and synthesis of such distinct luminophores, will assist in providing new effective turn-ON probes for non-invasive intravital chemiluminescence imaging techniques.