Alexa Fluor™ 647 Hydrazide - FAQs

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11 product FAQs found

我注入了荧光示踪剂，但在组织固定并切片后无法检测到它，哪个环节出了问题？

•请确认您所用的示踪剂交联到蛋白质上，或具有用于固定的伯胺——酰肼、赖氨酸可固定葡聚糖或蛋白质偶联物皆可。
•使用醛类固定剂交联示踪剂上的胺基。
•注入更大量或更高浓度的示踪剂。示踪剂通常注射1-20％浓度（10 mg/mL或更高）。
•确认您使用正确的荧光滤光片进行检测。您可吸取少量未稀释的示踪剂储液滴一滴到载玻片上，然后在显微镜下使用您所需的滤光片观察测试。这可验证示踪剂荧光是否可以被检测到，以及荧光显微镜的滤光片是否工作正常。
•回顾组织固定和处理步骤，确认是否存在可能影响示踪剂的试剂或处理步骤。

我用Alexa Fluor偶联生物胞素标记我的神经元以观察运输，但我想只检测逆向运输而生物胞素，似乎在同时逆向和顺向运输。 我该怎么办？

观察到这两种类型的运输是生物胞素的典型特征。带有标记的霍乱毒素B类的产品只进行逆向运输。

你们有类似于荧光黄（Lucifer Yellow）但颜色不同的神经元示踪剂吗？

Lucifer Yellow CH是一个酰肼，所以我们任一Alexa Fluor或荧光标记的酰肼产品都适于此用途。此类产品的清单请见此处（https://www.thermofisher.com/us/en/home/life-science/cell-analysis/cell-tracing-tracking-and-morphology/neuronal-tracing/hydrazides-biocytins.html#prd）。

我该如何为自己的试验选择示踪对象?

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）

你们有哪些神经元示踪产品？

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）。

What dye can I use that is non-reactive and can show an injection site?

The non-reactive Alexa Fluor and Alexa Fluor hydrazide derivatives may be used for injection site visualization. Other options include the fluorescent polystyrene microspheres, FluoSpheres, and dye-conjugated dextrans. The hydrazide derivatives and 'fixable' dextrans are retained by cross-linking using an aldehyde-based fixative.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

I injected a fluorescent tracer, but cannot detect it after tissue is fixed and sectioned. What am I doing wrong?

Confirm that the tracer you are using crosslinks to proteins or has a primary amine for fixation-either a hydrazide, lysine fixable dextran, or a protein conjugate.
Use aldehyde-based fixatives to cross link the amines on the tracer.
Inject a larger amount or higher concentration of the tracer. Tracers are generally injected at 1-20% concentrations (10 mg/mL or higher).
Confirm that you are using the correct fluorescent filter for detection. You can perform a spot test by pipetting a small amount of the undiluted stock solution of the tracer onto a slide, then view under the filter you are using on your microscope. This will confirm if the tracer fluorescence can be detected and the fluorescent microscope filter is working properly.
Review tissue fixation and handling procedures to confirm if any reagents or processing procedures could be affecting the tracer.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

I have labeled my neurons with an Alexa Fluor conjugated biocytin to look at transport but I wanted to examine only retrograde transport and biocytin appears to be moving retrograde and anterograde. What should I do?

Observing both types of transport is typical for biocytin. The conjugated cholera toxin subunit B products have been observed to travel only retrogradely.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

Do you have a neuronal tracer similar to Lucifer Yellow but in another fluorescent color?

Lucifer Yellow CH is a hydrazide, so any of our Alexa Fluor or fluorescent hydrazides could potentially be used. A listing of them can be found here. (https://www.thermofisher.com/us/en/home/life-science/cell-analysis/cell-tracing-tracking-and-morphology/neuronal-tracing/hydrazides-biocytins.html#prd)

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

How do I know which tracer to choose for my experiment?

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.

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.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.