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Based on a BODIPY molecular rotor and a zinc–dipicolylamine receptor, we designed a fluorogenic probe for the detection of apoptosis. Being poorly emissive in solution and with healthy cells, it selectively binds phosphatidylserine of early apoptotic cells and internalizes into late apoptotic cells, lighting up its green fluorescence. [ABSTRACT FROM AUTHOR]

Lipid droplets (LDs) are intracellular lipid-rich organelles that regulate the storage of neutral lipids and were recently found to be involved in many physiological processes, metabolic disorders, and diseases including obesity, diabetes, and cancers. Herein we present a family of new fluorogenic merocyanine fluorophores based on an indolenine moiety and a dioxaborine barbiturate derivative. These so-called StatoMerocyanines (SMCy) fluoresce from yellow to the near-infrared (NIR) in oil with an impressive fluorescence enhancement compared to aqueous media. Additionally, SMCy display remarkably high molar extinction coefficients (up to 390 000 M-1 cm-1) and high quantum yield values (up to 100%). All the members of this new family specifically stain the LDs in live cells with very low background noise. Unlike Nile Red, a well-known lipid droplet marker, SMCy dyes possess narrow absorption and emission bands in the visible, thus allowing multicolor imaging. SMCy proved to be compatible with fixation and led to high-quality 3D images of lipid droplets in cells and tissues. Their high brightness allowed efficient tissue imaging of adipocytes and circulating LDs. Moreover their remarkably high two-photon absorption cross-section, especially SMCy5.5 (up to 13 300 GM), as well as their capacity to efficiently fluoresce in the NIR region led to two-photon multicolor tissue imaging (liver). Taking advantage of the available color palette, lipid droplet exchange between cells was tracked and imaged, thus demonstrating intercellular communication. [ABSTRACT FROM AUTHOR]

We designed Calcium Rubies, a family of functionalizable BAPTA-based red-fluorescent calcium (Ca2+) indicators as new tools for biological Ca2+ imaging. The specificity of this Ca2+-indicator family is its side arm, attached on the ethylene glycol bridge that allows coupling the indicator to various groups while leaving open the possibility of aromatic substitutions on the BAPTA core for tuning the Ca2+-binding affinity. Using this possibility we now synthesize and characterize three different CaRubies with affinities between 3 and 22 μM. Their long excitation and emission wavelengths (peaks at 586/604 nm) allow their use in otherwise challenging multicolor experiments, e.g., when combining Ca2+ uncaging or optogenetic stimulation with Ca2+ imaging in cells expressing fluorescent proteins. We illustrate this capacity by the detection of Ca2+ transients evoked by blue light in cultured astrocytes expressing CatCh, a light-sensitive Ca2+-translocating channelrhodopsin linked to yellow fluorescent protein. Using time-correlated single-photon counting, we measured fluorescence lifetimes for all CaRubies and demonstrate a 10-fold increase in the average lifetime upon Ca2+ chelation. Since only the fluorescence quantum yield but not the absorbance of the CaRubies is Ca2+-dependent, calibrated two-photon fluorescence excitation measurements of absolute Ca2+ concentrations are feasible. [ABSTRACT FROM AUTHOR]

Nanoemulsions (NEs) are water-dispersed oil droplets that constitute stealth biocompatible nanomaterials. NEs can reach an impressive degree of fluorescent brightness owing to their oily core that can encapsulate a large number of fluorophores on the condition the latter are sufficiently hydrophobic and oil-soluble. BODIPYs are among the brightest green emitting fluorophores and as neutral molecules possess high lipophilicity. Herein, we synthesized three different natural lipid-BODIPY conjugates by esterification of an acidic BODIPY by natural lipids, namely: α-tocopherol (vitamin E), cholesterol, and stearyl alcohol. The new BODIPY conjugates were characterized in solvents and oils before being encapsulated in NEs at various concentrations. The physical (size, stability over time, leakage) and photophysical properties (absorption and emission wavelength, brightness, photostability) are reported and showed that the nature of the lipid anchor and the nature of the oil used for emulsification greatly influence the properties of the bright NEs. [ABSTRACT FROM AUTHOR]

Most chemical and, with only a few exceptions, all genetically encoded fluorimetric calcium (Ca 2+ ) indicators (GECIs) emit green fluorescence. Many of these probes are compatible with red-emitting cell- or organelle markers. But the bulk of available fluorescent-protein constructs and transgenic animals incorporate green or yellow fluorescent protein (GFP and YFP respectively). This is, in part, not only heritage from the tendency to aggregate of early-generation red-emitting FPs, and due to their complicated photochemistry, but also resulting from the compatibility of green-fluorescent probes with standard instrumentation readily available in most laboratories and core imaging facilities. Photochemical constraints like limited water solubility and low quantum yield have contributed to the relative paucity of red-emitting Ca 2+ probes compared to their green counterparts, too. The increasing use of GFP and GFP-based functional reporters, together with recent developments in optogenetics, photostimulation and super-resolution microscopies, has intensified the quest for red-emitting Ca 2+ probes. In response to this demand more red-emitting chemical and FP-based Ca 2+ -sensitive indicators have been developed since 2009 than in the thirty years before. In this topical review, we survey the physicochemical properties of these red-emitting Ca 2+ probes and discuss their utility for biological Ca 2+ imaging. Using the spectral separability index X ijk (Oheim M., 2010. Methods in Molecular Biology 591: 3–16) we evaluate their performance for multi-color excitation/emission experiments, involving the identification of morphological landmarks with GFP/YFP and detecting Ca 2+ -dependent fluorescence in the red spectral band. We also establish a catalog of criteria for evaluating Ca 2+ indicators that ideally should be made available for each probe. This article is part of a Special Issue entitled: Calcium signaling in health and disease. Guest Editors: Geert Bultynck, Jacques Haiech, Claus W. Heizmann, Joachim Krebs, and Marc Moreau. [ABSTRACT FROM AUTHOR]