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Fluorescent Cellular Stains

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Fluorescent Cellular Stains Membrane and Membrane Potential Dyes CellBrite™ Cytoplasmic Membrane Stains and Other Lipophilic Carbocyanine Dyes ... p. 2 Phospholipid Membrane Dyes ... p. 3 Membrane Potential Dyes ... p. 3 Synaptic Vesicle Dyes ... p. 4 Cytosolic Tracers ... p. 5 Organelle Stains MitoView™ Mitochondrial Dyes ... p. 6 LysoView™ Lysosome Stains ... p. 7 Cytoskeleton Probes ... p. 8 Nuclear Stains ... p. 9 Apoptosis and Viability Stains ... p. 10 Fluorescent Lectins, Toxins, and Other Conjugates ... p. 11 www.biotium.com • 1 Fluorescent Membrane and Membrane Potential Dyes CellBrite™ Cytoplasmic Membrane Dyes Lipophilic carbocyanine dyes are widely used for labeling neurons in tissues by retrograde labeling, and to label membranes in a wide variety of cell types. The dyes are weakly fluorescent in aqueous phase, but become highly fluorescent in lipid bilayers. Staining is highly stable with low toxicity and very little dye transfer in between cells, making the dyes suitable for long-term cell labeling and tracking studies. Cell populations can be labeled with different fluorescent colors for identification after mixing. Double labeling can identify cells that have fused or formed stable clusters. Unlike PKH dyes, CellBrite™ dyes do not require a complicated hypoosmotic labeling protocol. They are ready-to-use dye delivery solutions that can be added directly to normal culture media to label suspended or adherent cells in culture. We offer a selection of dyes with fluorescence ranging from blue to near-infrared. The CellBrite™ Blue Cytoplasmic Membrane Labeling Kit features DiB, a unique blue carbocyanine dye, and a cell loading agent. CellBrite™ Green (NeuroDiO), CellBrite™ Orange (DiI), CellBrite™ Red (DiD), and CellBrite™ NIR dyes are supplied as ready-to-add dye solutions. The near-infrared CellBrite™ NIR dyes are suitable for small animal imaging. Other Lipophilic Carbocyanine Dyes for Membrane Labeling Biotium also offers a selection of stand-alone carbocyanine dyes. DiOC14(3) is derivative of green fluorescent DiO that is more soluble in aqueous media. Neuro-DiO has improved solubility in membranes, less tendency to form non-fluorescent aggregates, and better photostability compared to DiO. Neuro-DiI is derivative of orange-fluorescent DiI with structural features to make the probe diffuse faster in cell membranes. Neuro-DiO and Figure 1. HeLa cells stained with 1 uM CellBrite™ NIR680 Neuro-DiI are available in vegetable oil, a formulation commonly used for microinjection. The (magenta) for 20 minutes at 37oC. Nuclei were stained with dilinoleyl carbocyanines (Fast DiI™ and Fast DiO™) have much faster lateral diffusion rate Hoechst (blue). than regular carbocyanine dyes and thus are excellent for labeling neurons in tissues. Near- infrared DiR can be used for small animal imaging. Absorption spectra of CellBrite™ Cytoplasmic Membrane Dyes Fluorescence emission spectra of CellBrite™ Cytoplasmic Membrane Dyes A CellBrite Blue B CellBrite Blue CellBrite Green CellBrite Green CellBrite Orange CellBrite Orange CellBrite Red CellBrite Red CellBrite NIR680 CellBrite NIR680 CellBrite NIR750 CellBrite NIR750 CellBrite NIR770 CellBrite NIR770 CellBrite NIR790 CellBrite NIR790 Emission Absorbance 250 350 450 550 650 750 850 375 475 575 675 775 875 Wavelength (nm) Wavelength (nm) Figure 2. Absorption (A) and emission (B) spectra of lipophilic carbocyanine dyes. CellBrite™ Blue and DiB have identical spectra. DiO, DiOC14(3), and Dilinoleyl DiO are spec- trally similar to CellBrite™ Green (Neuro-DiO). Neuro-DiI, and Dilinoleyl DiI are spectrally similar to CellBrite™ Orange (DiI). CellBrite™ Red and DiD have identical spectra. DiR is spectrally similar to CellBrite™ NIR750. CellBrite™ Cytoplasmic Membrane Dyes Other Lipophilic Carbocyanine Dyes Catalog Product Ex/Em (nm) Unit Size Catalog No. Product Ex/Em (nm) Unit Size No. 30024 CellBrite™ Blue Cytoplasmic Membrane Labeling Kit 366/441 50 assays 60011 DiO 484/501 50 mg 30021 CellBrite™ Green Cytoplasmic Membrane Labeling Dye 484/501 1 mL 60015 Neuro-DiO 484/501 25 mg 30022 CellBrite™ Orange Cytoplasmic Membrane Labeling Dye 549/565 1 mL 60019 Neuro-DiO in vegetable oil 484/501 200 uL 30023 CellBrite™ Red Cytoplasmic Membrane Labeling Dye 644/665 1 mL 60035 Dilinoleyl DiO (FAST DiO™) 484/499 5 mg 30070 CellBrite™ NIR680 Cytoplasmic Membrane Labeling Dye (2 mM) 683/724 100 uL 60012 DiOC14(3), hydroxyethanesulfonate 484/501 50 mg 30077 CellBrite™ NIR750 Cytoplasmic Membrane Labeling Dye (2 mM) 748/780 100 uL 60010 DiI 549/565 50 mg 30078 CellBrite™ NIR770 Cytoplasmic Membrane Labeling Dye (2 mM) 767/806 100 uL 60018 DiI in vegetable oil 549/565 0.5 mL 30079 CellBrite™ NIR790 Cytoplasmic Membrane Labeling Dye (2 mM) 786/820 100 uL 60034 Dilinoleyl DiI (FAST DiI™) 549/565 5 mg 60016 Neuro-DiI 549/565 25 mg 60014 DiD 644/655 50 mg 60017 DiR 748/780 25 mg Fast DiI and Fast DiO are trademarks of Thermo Fisher Scientific 2 • www.biotium.com Fluorescent Membrane and Membrane Potential Dyes Phospholipid Membrane Probes Fast-Responding Membrane Potential Dyes These membrane probes are derived from natural phospholipids by modifying the head Fast-responding membrane potential dyes are styryl dyes that undergo changes in group with a fluorescent dye or biotin. The probes are useful for studies of vesicle trafficking, fluorescence intensity in response to changes in membrane potential, on the order of 2-10% membrane fusion, and interaction of labeled membranes with other biomolecules. Red change in fluorescence per 100 mV. However, the dyes also undergo spectral shift with fluorescent phospholipids like TRITC-, Rhodamine-, and Texas Red®-DHPE have been changes in membrane potential, allowing ratiometric measurements. Fast response dyes used as fluorescence acceptors in combination with NBD-DHPE in FRET-based membrane- have been used to measure electrical activity in neural and cardiac cells. fusion assays. Di-4-ANEPPS and Di-8-ANEPPS show changes in fluorescence excitation intensity that correlate with changes in membrane potential. Di-8-ANNEPPS is more hydrophobic Catalog No. Product Ex/Em (nm) Unit Size and better retained in the outer leaflet of the plasma membrane than Di-4-ANNEPS, 60022 Biotin-DHPE N/A 10 mg and therefore is more suitable for long-term membrane potential studies. It is also more photostable and less phototoxic than Di-4-ANNEPS. 60023 Biotin-X-DHPE N/A 5 mg 60024 Fluorescein-DHPE 496/519 5 mg Di-2-ANEPEQ (also known as JPW 1114) is a highly water soluble fast-responding dye that is usually introduced into cells by microinjection. Di-8-ANEPQ and Di-12-ANEPQ are 60025 NBD-PE 463/536 10 mg successively more hydrophobic, and have been used for potential-sensitive retrograde 60028 TRITC-DHPE 540/566 1 mg labeling of neurons. 60026 Rhodamine-DHPE 560/581 5 mg RH237, RH414, RH421, and RH795 are fast-responding potentiometric probes generally 60027 Texas Red®-DHPE 582/601 1 mg used for functional imaging of neurons. RH421 exhibits >20% fluorescence change per 100 mV on neuroblastoma cells. These dyes can differ in their physiological effects, for example RH414 causes arterial constriction during cortex staining, while the spectrally similar dye Translational Membrane Potential Dyes RH795 does not. Translational (or slow-responding) membrane potential dyes undergo a change in their membrane distribution as a result of changes in membrane potential. DiBAC4(3) is a translational membrane potential dye that redistributes within the cell membrane when membrane potential changes. The fluorescence of the dye is enhanced with membrane depolarization. The rate of fluorescence response of the dye is slower than styryl dyes like the ANEPPS dyes, but the fluorescence signal change is significantly larger. Emission DiOC2(3) has been used for measuring membrane potential in bacteria. The green fluorescent dye forms red fluorescent aggregates with increasing membrane potential, Absorption allowing ratiometric potential measurements. DiOC5(3) and DiOC6(3) are two of the most widely used carbocyanine dyes for membrane potential measurements. 350 450 550 650 750 Wavelength (nm) Tetramethylrhodamine ethyl ester (TMRE) and Tetramethylrhodamine methyl ester (TMRM) can be used for quantitative measurements of membrane potentials using the Nernst Figure 1. Absorption and emission spectra of Di-4-ANEPPS and Di-8-ANEPPS equation. The dyes do not form aggregates in cell membranes and have minimal interaction bound to liposomes. The spectra of Di-2-ANEPEQ, Di-8-ANEPPQ, and Di-12- with membrane proteins. Thus, the transmembrane distribution of the dyes is directly ANEPPQ are expected to be similar but red-shifted by 10-20 nm. In general, the related to membrane potential. The dyes are also used as membrane potential-dependent spectra of styryl dyes undergo a large blue shift in going from organic solvent mitochondrial stains. to cell membranes. The spectra shown here should be representative of the spectra in cell membranes for ANEPPS and ANEPEQ dyes. Catalog No. Product Ex/Em (nm) (MeOH)* Unit Size 61010 Di-4-ANEPPS 496/705 5 mg 61012 Di-8-ANEPPS 498/713 5 mg Catalog No. Product Ex/Em (nm (MeOH) Unit Size 61013 Di-2-ANEPEQ (JPW 1114) see Fig. 1 5 mg 61011 DiBAC4(3) 493/516 25 mg 61014 Di-8-ANEPPQ see Fig. 1 5 mg 70008 DiOC2(3) 482/497 100 mg 61015 Di-12-ANEPPQ see Fig. 1 5 mg 70007 DiOC5(3) 482/497 100 mg 61018 RH237 528/782 5 mg 70009 DiOC6(3) 484/501 100 mg 61016 RH414 532/706 5 mg 70016 TMRE 549/574 25 mg 61017 RH421 515/704 25 mg 70017 TMRM 548/573 25 mg 61019 RH795 530/712 5 mg *Ex/Em is shown for methanol solution of dye. In cell membranes, spectra of styryl dyes are typically blue shifted by as much as 20 nm for absorption/excitation and 80 nm for emission Texas Red is a registered trademark of Thermo Fisher Scientific (see Fig. 1). www.biotium.com • 3 Fluorescent Membrane and Membrane Potential Dyes SynaptoGreen™ and SynaptoRed™ Nerve Terminal Dyes Nerve terminal probes were originally called FM® dyes.
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