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View additional product information for Dil Stain (1,1'-Dioctadecyl-3,3,3',3'-Tetramethylindocarbocyanine Perchlorate ('DiI'; DiIC18(3))) - FAQs (D282, D3911)
19 product FAQs found
如果您选择的示踪剂是亲脂性染料并且用甲醇固定，脂质会随着甲醇流失。如果您使用甲醇固定，可以选择能够共价结合神经元内的蛋白质的示踪剂。
由于这些染料插入脂质膜，任何对膜的破坏都将导致染料流失。这包括用Triton X-100之类的去垢剂或是甲醇之类的有机溶剂通透。通透是胞内抗体标记所必需的，但它会导致染料流失。相反，CFDA SE之类活性染料能够共价连接到细胞组分，因此可以在固定和通透之后更好的保留。
DiI是一种亲脂性染料，多数保留在细胞脂质内。当细胞经去垢剂通透或是用乙醇固定时，会去除掉脂质和染料。如果需要通透，可以使用CM-DiI标记，因为CM-DiI会共价结合膜里面的蛋白；部分信号会随着固定/通透丢失，但残留的信号仍足以被检测出。
转运过程相当缓慢：在活体组织中大约6 mm/天，在固定组织中更慢。亲脂性羰花青示踪剂从开始给样处扩散到神经元末端，可能需要数天甚至数周的时间。FAST DIO和DiI类似物（具有不饱和烷基尾）可以提高50％左右的转运速度。
请选择与您现有的激发光源和发射滤光片组/通道兼容的染料。固体、糊和晶体形式的染料可直接施加到组织中的神经元。如需标记培养的细胞或进行显微注射，可采用溶液或固态形式的亲脂性染料。
要考虑的因素有示踪对象的大小、给样方式（注射，直接上样到组织等），示踪对象是否需要固定。以下链接详细介绍了我们提供的各类神经元示踪剂的详细信息以及选择方法：
•神经元示踪（https://www.thermofisher.com/us/en/home/life-science/cell-analysis/cell-tracing-tracking-and-morphology/neuronal-tracing.html）
•选择示踪剂（https://www.thermofisher.com/us/en/home/references/molecular-probes-the-handbook/fluorescent-tracers-of-cell-morphology-and-fluid-flow/choosing-a-tracer.html）
•成像分析（http://assets.thermofisher.com/TFS-Assets/BID/Reference-Materials/bioprobes-50-journal.pdf）
固体和结晶形式的DiI以及其他相关染料（货号D282，D3911，D7757，和D12731）有时与一个特定的神经元接触，通过膜侧向扩散染上整个细胞。此外，我们的NeuroTrace组织标记糊剂也可挑到针上，置于特定的神经元上。 有关我们的神经元细胞标记方法的比较结果请参阅下表。 产品：标记方式：标记强度：特征 神经元特异性抗体：针对神经细胞表达蛋白的一抗：与蛋白质表达量成比例：唯一的神经元特异性标记方式。 亲脂性神经元示踪剂：疏水性染料掺入到细胞中的脂质：由于细胞中含有大量的脂类，这种标记方式能提供最强的标记：能够跟踪整个样品的神经元。 荧光标记的右旋糖苷：通常右旋糖苷采用显微注射：所有的细胞质可能被标记，标记密度可能会更高：能够标记单个神经元，从而避免了荧光背景干扰。 膜电位指示剂：染料通过水性缓冲液上样到活细胞里：取决于周围电场引起的结构变化，或者去极化引起的染料流入：膜电位的变化在各种生理进程中发挥核心作用，包括神经脉冲波传播，肌肉收缩以及细胞信号传导过程。
详细信息请查阅此网页（https://www.thermofisher.com/us/en/home/life-science/cell-analysis/cell-tracing-tracking-and-morphology/neuronal-tracing.html）。
亲脂性花青染料非常适合此类测定，因为它们会掺入到细胞膜中，随着细胞融合之后共用细胞膜，染料也随之共享。例如，一个细胞群体可以用DiI（橙红色）进行标记，而另一细胞群可以用DIO（绿色）进行标记，当细胞融合时合并的颜色为黄色（使用双带通滤光片成像）。
This is expected. DiD (which is far-red fluorescent) is never as uniform as DiI (which is orange fluorescent). If uniformity is desired, try increasing the label time and concentration, but it still isn't likely to be as uniform as DiI. CellMask Deep Red Plasma Membrane stain is much more uniform and is about the same wavelength as DiD. However, if you intend to do cell tracking over days, CellMask stain has not been tried for that application.
Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.
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.
Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.
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.
Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.
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.
Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.
The transport is fairly slow, around 6 mm/day in live tissue and slower in fixed tissue, so diffusion of lipophilic carbocyanine tracers from the point of their application to the terminus of a neuron can take several days to weeks The FAST DiO and DiI analogs (which have unsaturated alkyl tails) can improve transport rate by around 50%.
Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.
Select the dye that is compatible with your available excitation source(s) and emission filter set/channels. The solid, paste and crystal forms can be applied directly to neurons in tissues. For labeling cells in culture or microinjection, the lipophilic dyes in solution or solid form can be used.
Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.
Factors to consider are size of tracer, method of delivery (injection, direct application to tissue, etc.), and if the tracer needs to be fixable. Here are some links to details about the various classes of neuronal tracers we offer and how to choose between them:
Neuronal Tracing (https://www.thermofisher.com/us/en/home/life-science/cell-analysis/cell-tracing-tracking-and-morphology/neuronal-tracing.html)
Choosing a Tracer (https://www.thermofisher.com/us/en/home/references/molecular-probes-the-handbook/fluorescent-tracers-of-cell-morphology-and-fluid-flow/choosing-a-tracer.html)
Imaging Analysis (http://assets.thermofisher.com/TFS-Assets/BID/Reference-Materials/bioprobes-50-journal.pdf)
Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.
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
Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.
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.
Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.
Lipophilic cyanine dyes are preferred for this sort of assay, since they insert into cellular membranes and then, upon fusion, are shared by the fused cells as the membranes are shared. For example, one cell population can be labeled with DiI (orange-red) and another cell population can be labeled with DiO (green), and when the cells fuse, the combined color appears yellow (when imaged with a dual-bandpass filter set).
Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.