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Invitrogen™ Dextran, Tetramethylrhodamine and biotin, 10,000 MW, Lysine Fixable (mini-Ruby)

Catalog No. D3312
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D3312 10 mg
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Catalog No. D3312 Supplier Invitrogen™ Supplier No. D3312
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Description

Rigorous methods for removing as much unconjugated dye as practical, followed by assay of dextran conjugates by thin-layer chromatography to help ensure absence of low molecular weight contaminants

Labeled dextrans are hydrophilic polysaccharides most commonly used in microscopy studies to monitor cell division, track the movement of live cells, and to report the hydrodynamic properties of the cytoplasmic matrix. The labeled dextran is commonly introduced into the cells via microinjection. Dextrans are hydrophilic polysaccharides characterized by their moderate-to-high molecular weight, good water solubility, and low toxicity. They also generally exhibit low immunogeniticy. Dextrans are biologically inert due to their uncommon poly-(α-D-1,6-glucose) linkages, which render them resistant to cleavage by most endogenous cellular glycosidases.

  • Succinimidyl coupling of dyes to dextran molecule, which, in most cases, results in neutral or anionic dextran
  • Reaction used to produce Rhodamine Green™ and Alexa Fluor™ 488 dextrans results in final product being neutral, anionic, or cationic
  • Suitable for neuronal tracing (anterograde and retrograde) in live cells
  • Cell lineage tracing in live cells
  • Examines intercellular communications (e.g., in gap junctions, during wound healing, and during embryonic development)
  • Investigates vascular permeability and blood-brain barrier integrity
  • Monitors acidification (some dextran-dye conjugates are pH-sensitive)
  • Studies hydrodynamic properties of cytoplasmic matrix

  • Most of dextrans labeled with tetramethylrhodamine dyes are essentially neutral
  • To produce more highly anionic dextrans, proprietary procedure adds negatively charged groups to dextran carriers; these products are designated polyanionic dextrans
  • Some applications require that dextran tracer be treated with formaldehyde or glutaraldehyde for subsequent analysis
  • For these applications, lysine-fixable dextran conjugates of fluorophores or biotin
  • Dextrans have covalently bound lysine residues that permit dextran tracers to be conjugated to surrounding biomolecules by aldehyde-mediated fixation for subsequent detection by immunohistochemical and ultrastructural techniques

Cell Analysis, Cell Tracing and Tracking, General Cell Tracing, Neuronal Tracing

Order Info

Shipping Condition: Room temperature

Specifications

Product Type Dextran
Quantity 10 mg
Content And Storage Store in freezer (-5 to -30°C) and protect from light.
Label or Dye Classic Dyes
Shipping Condition Room Temperature
Excitation/Emission 555/580 nm
Product Line Invitrogen

Frequently Asked Questions (FAQs)

I can't see the structural details of neurons when I inject my fluorescent dextran. What can I do to improve the detailed structure?

If you want to see the most detailed structure you should use the low molecular weight conjugated dextrans such as the 3,000 MW dextrans.
Why isn't my fluorescently conjugated dextran signal retained after fixation?

Ensure that the dextran you are using is the fixable form (i.e., contains a primary amine). Dextrans that do not contain a primary amine will not be fixed. Another factor could be that the concentration of the dextran is too low, and the concentration use can be increased up to 10 mg/mL.
Why do I lose all signal from my neuronal tracer when I do a methanol fixation on my cells?

If the tracer you chose is a lipophilic dye and fix with methanol, the lipids are lost with the methanol. If you have to use methanol fixation then choose a tracer that will covalently bind to proteins in the neurons.
I stained my cells with a lipophilic cyanine dye, like DiI, but the signal was lost when I tried to follow up with antibody labeling. Why?

Since these dyes insert into lipid membranes, any disruption of the membranes leads to loss of the dye. This includes permeabilization with detergents like Triton X-100 or organic solvents like methanol. Permeabilization is necessary for intracellular antibody labeling, leading to loss of the dye. Instead, a reactive dye such as CFDA SE should be used to allow for covalent attachment to cellular components, thus providing for better retention upon fixation and permeabilization.
I labeled my neurons with DiI and then fixed and permeabilized and now I have no signal. What did I do wrong?

DiI is a lipophilic dye that resides mostly in lipids in the cell, when cells are permeabilized with detergent or fixed using alcohol this strips away the lipid and the dye. If permeabilization is required CM-DiI can be used because this binds covalently to proteins in the membrane; some signal is lost upon fixation/permeabilization, but enough signal should be retained to make detection possible.
What are the charges of the dextrans?

We do not determine the net charge of the dextran conjugates. The net charge depends on the fluorophore used to label the dextran and the method of preparing the conjugate. We label some dextrans as neutral or anionic based on the fluorophore used, however the net charge of the dextran may not always be the same as the dye. The Alexa Fluor, Cascade Blue, Lucifer Yellow, fluorescein, and Oregon Green dextrans are intrinsically anionic, whereas most of the dextrans labeled with the zwitterionic Rhodamine B, tetramethylrhodamine and Texas Red dyes are essentially neutral.
What size dextran is best for neuronal tracing?

Dextrans with molecular weights from 3,000 to 70,000 have been used, however the 3,000 and 10,000 MW dextrans are most commonly used for neuronal tracing. The 3,000 MW dextrans are used for more detailed tracing of fine neuronal projections, investigating gap junctions, and diffuse more quickly; while the 10,000 MW dextrans have slower distribution, longer cellular retention, and do not cross gap junctions.
Do you have a neuronal tracing protocol?

The NeuroTrace BDA-10,000 Neuronal Tracer Kit (Cat. No. N7167) manual has a good protocol for injection procedures and neuronal tracing using the10,000 MW lysine-fixable biotin dextran amine (BDA). This protocol could potentially be applied to other fluorescent dextrans.

Please review Tables 1a and 1b on pages 4 and 5 - https://tools.thermofisher.com/content/sfs/manuals/mp07167.pdf
Is there a way to label individual neurons without microinjecting?

The solid and crystalline forms of DiI and other related dyes (Cat. Nos. D282, D3911, D7757, and D12731) are sometimes placed in contact with a specific neuron where it will travel down the cell by lateral diffusion via the membrane. Alternatively, our NeuroTrace Tissue Labeling Paste can be scooped onto a needle and placed onto particular neurons.

Please see the information below for a comparison of our neuronal cell labeling methods:
Product:Method of labeling: Labeling intensity: Features
Neuron-specific antibodies: Primary antibodies directed to proteins expressed in neuronal cells: Proportional to the amount of protein expressed: Provides the only neuronal specific labeling method
Lipophilic neuronal ytracers: Hydrophobic dyes are incorporated into lipids in the cell: This labeling method provides the most intense labeling becuase of the abundant amount of lipids: Allows tracing of neurons throughout the sample
Membrane potential indicators: Dyes are loaded into live cells in aqueous buffers: Depends on either changes in structures due to the electrical field they are in, or dye influx due to depolarization: Changes in membrane potential play a central role in physiological processes, including nerve-impulse propagation, muscle contraction, and cell signaling
What products do you have for neuronal tracing?

Please check out this web page (https://www.thermofisher.com/us/en/home/life-science/cell-analysis/cell-tracing-tracking-and-morphology/neuronal-tracing.html) for details.

For Research Use Only. Not for use in diagnostic procedures.