Abstract: High aldehyde dehydrogenase 1A1 (ALDH1A1) activity has emerged as a reliable marker for the identification of both normal and cancer stem cells. Herein, is presented AlDeSense, a turn-on green fluorescent probe for aldehyde dehydrogenase 1A1 (ALDH1A1) and Ctrl-AlDeSense, a matching non-responsive reagent. AlDeSense exhibits a 20-fold fluorescent enhancement when treated with ALDH1A1. Through the application of surface marker antibody staining, tumorsphere assays, and assessment of tumorigenicity, the disclosed results show that cells exhibiting high AlDeSense signal intensity have properties of cancer stem cells. Herein, is also reported the development of a red congener, red-AlDeSense. Importantly, red-AlDeSense represents one of only a few examples of a turn-on sensor in the red region using the d-PeT quenching mechanism.

Abstract: Detection of nitroxyl (HNO), the transient one-electron reduced form of nitric oxide, is a significant challenge owing to its high reactivity with biological thiols (rate constants as high as 109M?1 s?1). Reported herein is a new thiol-based HNO-responsive trigger that can compete against reactive thiols for HNO. This process forms an N-hydroxysulfenamide intermediate which cyclizes to release a masked fluorophore leading to fluorescence enhancement. To ensure a rapid cyclization step, the disclosed design capitalizes on two established physical organic phenomena: the alpha-effect and the Thorpe-Ingold effect. Using this new trigger, NitroxylFluor was developed; a selective HNO-responsive fluorescent probe. Treatment of NitroxylFluor with an HNO donor results in a 16-fold turn-on. This probe also exhibits excellent selectivity over various reactive nitrogen, oxygen, and sulfur species and efficacy in the presence of thiols (e.g., glutathione in mM concentrations).

Abstract: Hypoxia occurs when limited oxygen supply impairs physiological functions and is a pathological hallmark of many diseases including cancer and ischemia. Thus, detection of hypoxia can guide treatment planning and serve as a predictor of patient prognosis. Current methods suffer from invasiveness, poor resolution and low specificity. To address these limitations, various hypoxia-responsive probes (HyPs) for photoacoustic imaging are disclosed. The emerging modality converts safe, non-ionizing light to ultrasound waves, enabling acquisition of high-resolution 3D images in deep tissue. The HyPs feature an N-oxide trigger that is reduced in the absence of oxygen by haem proteins such as CYP450 enzymes. Reduction of HyPs produce a spectrally distinct product, facilitating identification via photoacoustic imaging. HyPs exhibit selectivity for hypoxic activation in vitro, in living cells and in multiple disease models in vivo.

Abstract: Cyclooxygenase-2 (COX-2) over-expression is prominent in inflammatory diseases, neurodegenerative disorders, and cancer. Directly monitoring COX-2 activity within its native environment poses an exciting approach to account for and illuminate the effect of the local environments on protein activity. Herein, we report the development of CoxFluor, the first activity-based sensing approach for monitoring COX-2 within live cells with confocal microscopy and flow cytometry. CoxFluor strategically links a natural substrate with a dye precursor to engage both the cyclooxygenase and peroxidase activities of COX-2. This catalyzes the release of resorufin and the natural product, as supported by molecular dynamics and ensemble docking. CoxFluor enabled the detection of oxygen-dependent changes in COX-2 activity that are independent of protein expression within live macrophage cells.

Abstract: Probes for formaldehyde (FA) including a homoallylamine trigger group attached to a detectable moiety are provided. Aspects of the probes include luminogenic or fluorogenic probes, such as a probe including a quencher in energy-receiving proximity to a fluorophore. Also provided are methods of using the probes for sensitive and bio orthogonal detection of FA in a sample. Aspects of the methods include selectively reacting the probe with the formaldehyde in the sample to a release (e.g., via a 2-aza-Cope rearrangement) a reporter group comprising a detectable moiety. Aspects of the methods detecting formaldehyde in a cell, tissue, organ or fluid in a subject. Also provided are compositions and kits including the subject probes that find use in practicing various embodiments of the subject methods.

Abstract: High aldehyde dehydrogenase 1A1 (ALDH1A1) activity has emerged as a reliable marker for the identification of both normal and cancer stem cells. Herein, is presented AlDeSense, a turn-on green fluorescent probe for aldehyde dehydrogenase 1A1 (ALDH1A1) and Ctrl-AlDeSense, a matching non-responsive reagent. AlDeSense exhibits a 20-fold fluorescent enhancement when treated with ALDH1A1. Through the application of surface marker antibody staining, tumorsphere assays, and assessment of tumorigenicity, the disclosed results show that cells exhibiting high AlDeSense signal intensity have properties of cancer stem cells. Herein, is also reported the development of a red congener, red-AlDeSense. Importantly, red-AlDeSense represents one of only a few examples of a turn-on sensor in the red region using the d-PeT quenching mechanism.

Abstract: Detection of nitroxyl (HNO), the transient one-electron reduced form of nitric oxide, is a significant challenge owing to its high reactivity with biological thiols (rate constants as high as 109M?1 s?1). Reported herein is a new thiol-based HNO-responsive trigger that can compete against reactive thiols for HNO. This process forms an N-hydroxysulfenamide intermediate which cyclizes to release a masked fluorophore leading to fluorescence enhancement. To ensure a rapid cyclization step, the disclosed design capitalizes on two established physical organic phenomena: the alpha-effect and the Thorpe-Ingold effect. Using this new trigger, NitroxylFluor was developed; a selective HNO-responsive fluorescent probe. Treatment of NitroxylFluor with an HNO donor results in a 16-fold turn-on. This probe also exhibits excellent selectivity over various reactive nitrogen, oxygen, and sulfur species and efficacy in the presence of thiols (e.g., glutathione in mM concentrations).

Abstract: Hypoxia occurs when limited oxygen supply impairs physiological functions and is a pathological hallmark of many diseases including cancer and ischemia. Thus, detection of hypoxia can guide treatment planning and serve as a predictor of patient prognosis. Current methods suffer from invasiveness, poor resolution and low specificity. To address these limitations, various hypoxia-responsive probes (HyPs) for photoacoustic imaging are disclosed. The emerging modality converts safe, non-ionizing light to ultrasound waves, enabling acquisition of high-resolution 3D images in deep tissue. The HyPs feature an N-oxide trigger that is reduced in the absence of oxygen by haem proteins such as CYP450 enzymes. Reduction of HyPs produce a spectrally distinct product, facilitating identification via photoacoustic imaging. HyPs exhibit selectivity for hypoxic activation in vitro, in living cells and in multiple disease models in vivo.

Abstract: Probes for formaldehyde (FA) including a homoallylamine trigger group attached to a detectable moiety are provided. Aspects of the probes include luminogenic or fluorogenic probes, such as a probe including a quencher in energy-receiving proximity to a fluroophore. Also provided are methods of using the probes for sensitive and bio orthogonal detection of FA in a sample. Aspects of the methods include selectively reacting the probe with the formaldehyde in the sample to a release (e.g., via a 2-aza-Cope rearrangement) a reporter group comprising a detectable moiety. Aspects of the methods detecting formaldehyde in a cell, tissue, organ or fluid in a subject. Also provided are compositions and kits including the subject probes that find use in practicing various embodiments of the subject methods.

Abstract: The invention provides fluorescent sensors for the selective detection of a metal such as copper. The sensors may be considered to be derivatives of cyanine, fluorescein, rhodamine, rhodol, Tokyo green, or BODIPY. The sensors find particular use in detecting copper in cells and living animals.