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Experimental Approaches to the Study of A2E, a Bisretinoid Lipofuscin Chromophore of Retinal Pigment Epithelium

Bisretinoid lipofuscin compounds that accumulate in retinal pigment epithelial (RPE) cells are implicated in the pathogenesis of some forms of macular degeneration. In the development of approaches to the amelioration of retinal disorders characterized by enhanced RPE lipofu ...

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Culture of Highly Differentiated Human Retinal Pigment Epithelium for Analysis of the Polarized Uptake, Processing, and Secretion of Retinoids

The retinal pigment epithelium (RPE) occupies a strategic position within the eye, given its location between the neurosensory retina and the vascular bed (choroid) that nourishes the photoreceptor cells (rods and cones). Among the many attributes of this versatile monolayer of cells is ...

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Feeder-Independent Culture of Mouse Embryonic Stem Cells Using Vitamin A/Retinol

Embryonic stem (ES) cells derived from the inner cell mass of a mammalian blastocyst represent unlimited source of all types of cells for regenerative medicine and for drug discovery. Mouse and human ES cells require mouse embryonic fibroblast feeder cells to maintain their undifferenti ...

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Techniques to Study Specific Cell-Surface Receptor-Mediated Cellular Vitamin A Uptake

STRA6 is a multitransmembrane domain protein that was recently identified as the cell-surface receptor for plasma retinol-binding protein (RBP), the vitamin A carrier protein in the blood. STRA6 binds to RBP with high affinity and mediates cellular uptake of vitamin A from RBP. It is not homolo ...

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Quantification of Endogenous Retinoids

Numerous physiological processes require retinoids, including development, nervous system function, immune responsiveness, proliferation, differentiation, and all aspects of reproduction. Reliable retinoid quantification requires suitable handling and, in so ...

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Binding of Retinoids to ABCA4, the Photoreceptor ABC Transporter Associated with Stargardt Macular Degeneration

ABCA4 is a member of the superfamily of ATP-binding cassette (ABC) transporters, which has been implicated in the clearance of all-trans retinal derivatives from rod and cone photoreceptor cells following photoexcitation as part of the visual cycle. Mutations in ABCA4 are known to cause St ...

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Microfluorometric Measurement of the Formation of All-Trans-Retinol in the Outer Segments of Single Isolated Vertebrate Photoreceptors

The first step in the detection of light by vertebrate photoreceptors is the photoisomerization of the retinyl chromophore of their visual pigment from 11-cis to the all-trans configuration. This initial reaction leads not only to an activated form of the visual pigment, meta II, that initia ...

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Measurement of the Mobility of All-Trans-Retinol with Two-Photon Fluorescence Recovery After Photobleaching

The mobility of all-trans-retinol makes a crucial contribution to the rate of the reactions in which it participates. This is even more so because of its low aqueous solubility, which makes the presence of carrier proteins and the spatial arrangement of cellular membranes especially releva ...

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Physiological Studies of the Interaction Between Opsin and Chromophore in Rod and Cone Visual Pigments

The visual pigment in vertebrate photoreceptors is a G protein-coupled receptor that consists of a protein, opsin, covalently attached to a chromophore, 11-cis-retinal. Activation of the visual pigment by light triggers a transduction cascade that produces experimentally measu ...

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In Vitro Assays of Rod and Cone Opsin Activity: Retinoid Analogs as Agonists and Inverse Agonists

Upon absorption of a photon, the bound 11-cis-retinoid isomerizes to the all-trans form resulting in a protein conformational change that enables it to activate its G protein, transducin, to begin the visual signal transduction cascade. The native ligand, 11-cis-retinal, acts as an inverse ...

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HPLC/MSN Analysis of Retinoids

This protocol describes a highly sensitive and selective method to quantify retinoids using normal-phase HPLC with online APCI MSN. The retinoids are key regulators of gene expression, retinol being oxidized via a retinaldehyde intermediate to retinoic acid (RA) which activates spe ...

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Enantioseparations of Primary Amino Compounds by High-Performance Liquid Chromatography Using Chiral Crown Ether-Based Chiral Stationary Phase

Liquid chromatographic resolution of racemic compounds containing a primary amino group has been known to be most successful when chiral crown ether-based chiral stationary phases (CSPs) are used. Among various crown ether-based CSPs, the stationary phase based on (+)-(18-crown-6)-2 ...

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Enantioseparations by High-Performance Liquid Chromatography Using Macrocyclic Glycopeptide-Based Chiral Stationary Phases: An Overview

Since their introduction by Armstrong in 1994, macrocyclic antibiotic-based chiral stationary phases have proven their applicability for the chiral resolution of various types of racemates. The unique structure of macrocyclic glycopeptides and their large variety of intera ...

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Chiral Separations by HPLC on Immobilized Polysaccharide Chiral Stationary Phases

Immobilized polysaccharide chiral stationary phases (CSPs) are a new development in chiral chromatography with increasing importance for the resolution of racemic compounds. These CSPs can be used with solvents traditionally applied for the classical coated polysaccharide ...

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Common Screening Approaches for Efficient Analytical Method Development in LC and SFC on Columns Packed with Immobilized Polysaccharide-Derived Chiral

Owing to their remarkable enantioselectivity, versatility, and stability, immobilized polysaccharide-based chiral stationary phases (CSPs) have been successfully integrated into the tool box of many research and industry groups for the separation of enantiomers or stere ...

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Enantioseparations by High-Performance Liquid Chromatography Using Polysaccharide-Based Chiral Stationary Phases: An Overview

This chapter summarizes the application of polysaccharide-based chiral stationary phases (CSPs) for separation of enantiomers in high-performance liquid chromatography (HPLC). Since this book contains dedicated chapters on enantioseparations using supercritical ...

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HPLC Enantioseparation on Cyclodextrin-Based Chiral Stationary Phases

Cyclodextrin-bonded silica material is one of the most commonly used chiral stationary phases in liquid chromatography for pharmaceutical analysis and enantioseparations. The approaches for immobilization of cyclodextrins onto the silica surface influence both, the sta ...

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Gas-Chromatographic Enantioseparation of Unfunctionalized Chiral Hydrocarbons: An Overview

Analytical gas-chromatographic enantioseparations of unfunctionalized chiral hydrocarbons (cycloalkanes and alkanes) on modified cyclodextrins with high-resolution capillary columns are reviewed. Due to the lack of functional groups of the hydrocarbons, enanti ...

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Cyclodextrin-Mediated Enantioseparations by Capillary Electrochromatography

Immobilized cyclodextrin derivatives are used as chiral selectors in various modes of capillary electrochromatography (CEC). The present chapter describes three techniques in detail utilizing CDs in CEC: (1) open-tubular capillary electrochromatography (o-CEC), (2) pack ...

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Enantioseparations in Capillary Electrochromatography Using Sulfated Poly -Cyclodextrin-Modified Silica-Based Monolith as Stationary Phase

As the most popular chiral selectors, β-cyclodextrin (β-CD) and its derivatives have been widely used in enantiomer separations. Among functionalized β-CDs, sulfated β-CDs are recognized as the most effective chiral additives in capillary electrophoresis (CE) separations. Here ...

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