Defining the Basis of Cyanine Phototruncation Enables a New Approach to Single-Molecule Localization Microscopy

The light-promoted conversion of extensively used cyanine dyes to blue-shifted emissive products has been observed in various contexts. However, both the underlying mechanism and the species involved in this photoconversion reaction have remained elusive. Here we report that irradiation of heptamethine cyanines provides pentamethine cyanines, which, in turn, are photoconverted to trimethine cyanines. We detail an examination of the mechanism and substrate scope of this remarkable twocarbon phototruncation reaction. Supported by computational analysis, we propose that this reaction involves a singlet oxygeninitiated multistep sequence involving a key hydroperoxycyclobutanol intermediate. Building on this mechanistic framework, we identify conditions to improve the yield of photoconversion by over an order of magnitude. We then demonstrate that cyanine phototruncation can be applied to super-resolution single-molecule localization microscopy, leading to improved spatial resolution with shorter imaging times. We anticipate these insights will help transform a common, but previously mechanistically ill-defined, chemical transformation into a valuable optical tool

Instrumental and sensory approaches for the characterization of compounds responsible for wine aroma

More than 800 aromatic compounds have been identified in wine, some of them at the ng/l level. Wine, therefore, constitutes a very complex matrix, from which it is difficult to isolate a specific aroma character. Gas chromatography–olfactometry (GC–O) applied to wine extracts is used to characterize odor-active zones that are often treated in a hierarchical way by Aroma Extract Dilution Analysis (AEDA). The aromatic impact of the volatiles is evaluated, generally by determining perception thresholds. This methodology has provided convincing results concerning wine flavors, but it does have its limitations. Forinstance , data on b-damascenone have demonstrated that these methods could reach their limits for this volatile, in particular, because of the non-quantitative representation of aroma extracts of wines, and because of the difficulty to accurately determine the perception threshold in wines for a compound already present. For b-damascenone, we have shown that its very low detection threshold with GC–O, its wide range, and its dependence on the composition of the medium resulted in overestimating its direct impact on the aroma of wine. Another way to facilitate the characterization of aromatic compounds was, therefore, investigated. High-Performance Liquid Chromatography (HPLC) methods were developed for the analysis of wine extracts. From an aromatic extract, 25 fractions with various flavors were thus obtained, and reverse-phase methodology was used for the selection and characterization of red- and black-fruit aromas in red wines

High-Contrast Detection of Somatostatin Receptor Subtype-2 for Fluorescence-Guided Surgery

Dye design can influence the ability of fluorescently labeled imaging agents to generate tumor contrast and has become an area of significant interest in the field of fluorescence-guided surgery (FGS). Here, we show that the charge-balanced near-infrared fluorescent (NIRF) dye FNIR-Tag enhances the imaging properties of a fluorescently labeled somatostatin analogue

The E-NTPDase family of ectonucleotidases: Structure function relationships and pathophysiological significance

Ectonucleotidases are ectoenzymes that hydrolyze extracellular nucleotides to the respective nucleosides. Within the past decade, ectonucleotidases belonging to several enzyme families have been discovered, cloned and characterized. In this article, we specifically address the cell surface-located members of the ecto-nucleoside triphosphate diphosphohydrolase (E-NTPDase/CD39) family (NTPDase1,2,3, and 8). The molecular identification of individual NTPDase subtypes, genetic engineering, mutational analyses, and the generation of subtype-specific antibodies have resulted in considerable insights into enzyme structure and function. These advances also allow definition of physiological and patho-physiological implications of NTPDases in a considerable variety of tissues. Biological actions of NTPDases are a consequence (at least in part) of the regulated phosphohydrolytic activity on extracellular nucleotides and consequent effects on P2-receptor signaling. It further appears that the spatial and temporal expression of NTPDases by various cell types within the vasculature, the nervous tissues and other tissues impacts on several patho-physiological processes. Examples include acute effects on cellular metabolism, adhesion, activation and migration with other protracted impacts upon developmental responses, inclusive of cellular proliferation, differentiation and apoptosis, as seen with atherosclerosis, degenerative neurological diseases and immune rejection of transplanted organs and cells. Future clinical applications are expected to involve the development of new therapeutic strategies for transplantation and various inflammatory cardiovascular, gastrointestinal and neurological diseases

Alpha-Ammonium Carbamates Undergo Efficient Two-Step Linker Cleavage and Improve the Properties of Antibody Conjugates

Targeted payload delivery strategies, such as antibody-drug conjugates (ADCs), have emerged as useful therapeutics. Although considerable efforts have been made in the areas of antibody engineering and labeling methodology, improving the overall physicochemical properties of the linker/payload combination remains an important challenge. Here we report an approach to create an intrinsically polar linker domain. We find that -ammonium carbamates (AAC) undergo a tandem 1,6–1,2 elimination sequence to release secondary amines. Using a fluorogenic hemicyanine as a model payload component, we show that a zwitterionic AAC linker improves labeling efficiency and reduces antibody aggregation when compared to a commonly used Val-Ala linker as well as a cationic AAC. Cellular and in vivo fluorescence imaging studies demonstrate that the model payload is released in antigen-expressing cells and tumors with high specificity. Broadly, this strategy may provide a general approach to mask the hydrophobicity of payloads and improve the properties of targeted agents

Ureter Identification In an Inflammatory Pig Model Using a Novel Near‐Infrared Fluorescent Dye

BACKGROUND AND OBJECTIVES: Ureters are at risk of injury in settings of inflammation and distorted anatomy. The use of a fluorescent dye can improve intraoperative ureteral identification without the need for any additional invasive procedures. Our team has previously described the development of a preclinical ureter-specific dye, UL-766, tested in a rat model. Here, we present the use of the fluorescent dye during laparoscopy to assist in ureteral identification in a swine model with an inflamed abdomen; the results of this study serve as proof of feasibility for use in the setting of tissue edema and erythema. STUDY DESIGN/MATERIALS AND METHODS: With institutional approval, two 20–25 kg pigs underwent abdominal surgery with the use of a Food and Drug Administration-approved fluorescence laparoscopic system. Using standard laparoscopy, inflammation was induced with sharp and blunt dissection and irritation was induced with gauze. The animals were allowed to recover and returned to the operating room after 7 days. Images of the inflamed right retroperitoneum, with fluorescence imaging, turned on, were taken before and after intravenous injection of the novel fluorescent dye at 120 μg/kg. The time until fluorescence visualization of the ureters was measured, and the fluorescent signal was measured for up to 4 hours from the time of the initial dye injection. Partial and complete transection of ureteral injuries was made by scissors and monitored under both standard video and fluorescence laparoscopy. RESULTS: Inflammation reduced the certainty of ureter identification by white light alone. Despite surrounding tissue erythema and edema, ureteral visualization under fluorescence laparoscopy was achieved within 5–10 minutes after dye injection. The fluorescent signal remained visible for at least 4 hours after injection, and the fluorescent dye showed a partial ureteral injury that would not have been observed under standard laparoscopy. CONCLUSIONS: UL-766 is a preclinical fluorescent dye useful for the intraoperative identification of the ureters and ureteral injuries in an inflamed abdomen. With the acquisition of additional preclinical data, this novel dye can be a valuable tool during laparoscopic abdominal and pelvic surgeries. Lasers Surg. Med. © 2019 Wiley Periodicals, Inc

Ureter Identification In an Inflammatory Pig Model Using a Novel Near-Infrared Fluorescent Dye.

BACKGROUND AND OBJECTIVES: Ureters are at risk of injury in settings of inflammation and distorted anatomy. The use of a fluorescent dye can improve intraoperative ureteral identification without the need for any additional invasive procedures. Our team has previously described the development of a preclinical ureter-specific dye, UL-766, tested in a rat model. Here, we present the use of the fluorescent dye during laparoscopy to assist in ureteral identification in a swine model with an inflamed abdomen; the results of this study serve as proof of feasibility for use in the setting of tissue edema and erythema. STUDY DESIGN/MATERIALS AND METHODS: With institutional approval, two 20–25 kg pigs underwent abdominal surgery with the use of a Food and Drug Administration-approved fluorescence laparoscopic system. Using standard laparoscopy, inflammation was induced with sharp and blunt dissection and irritation was induced with gauze. The animals were allowed to recover and returned to the operating room after 7 days. Images of the inflamed right retroperitoneum, with fluorescence imaging, turned on, were taken before and after intravenous injection of the novel fluorescent dye at 120 μg/kg. The time until fluorescence visualization of the ureters was measured, and the fluorescent signal was measured for up to 4 hours from the time of the initial dye injection. Partial and complete transection of ureteral injuries was made by scissors and monitored under both standard video and fluorescence laparoscopy. RESULTS: Inflammation reduced the certainty of ureter identification by white light alone. Despite surrounding tissue erythema and edema, ureteral visualization under fluorescence laparoscopy was achieved within 5–10 minutes after dye injection. The fluorescent signal remained visible for at least 4 hours after injection, and the fluorescent dye showed a partial ureteral injury that would not have been observed under standard laparoscopy. CONCLUSIONS: UL-766 is a preclinical fluorescent dye useful for the intraoperative identification of the ureters and ureteral injuries in an inflamed abdomen. With the acquisition of additional preclinical data, this novel dye can be a valuable tool during laparoscopic abdominal and pelvic surgeries. Lasers Surg. Med. © 2019 Wiley Periodicals, Inc