UltraComp eBeads™ Compensation Beads, 100 tests - FAQs

View additional product information for UltraComp eBeads™ Compensation Beads - FAQs (01-2222-42, 01-2222-41)

12 product FAQs found

我在进行流式细胞实验时，补偿值超过100，可能出现这种情况吗？

这是可能的。但是如果您的值超过100，可能需要查看你们用来采集数据的电压。

有人告诉我说我需要补偿自己的流式细胞分析数据，这是什么意思？

通过运行单色对照，有可能除去溢出到另一个荧光采集通道的荧光信号。

流式细胞分析该选用哪种类型的对照？

进行补偿时，你需要为自己使用的每种颜色参数准备单一染色样品（或补偿微球）。此外，我们建议您使用FMO（荧光扣除）对照。您可以使用减去一种颜色的其他所有颜色标记您的细胞或者微球来作为对照。为各种颜色准备FMO对照。这些对照对正确设置数据门限很有帮助。

在流式细胞分析中为何要担心补偿？

理想情况下，每一个单独荧光团的荧光发射光谱都非常集中，窄峰，发射峰之间分隔的较开。现实中，有机染料和荧光蛋白有很宽的发射峰，必须考虑补偿（数据采集中或后）来矫正任何一个荧光团荧光信号的分配。补偿从重叠光谱中移除了荧光信号，使您知道所看见的信号就是目标荧光团发出的信号，所以非常重要。

如何为抗体设置补偿对照？

补偿对照仅需未染色的细胞样品（或阴性对照）和单一颜色标记的样品——每管一个。如果您希望单色染色对照如您期望的那么亮,，那么可以使用我们的AbC总抗体补偿微珠试剂盒（货号A10513和A10497），这试剂盒是将抗体结合到微珠而非细胞上。

How long are UltraComp eBeads Compensation Beads (Cat. No. 01-2222-41, 01-2222-42) stable after staining?

After staining, UltraComp eBeads Compensation Beads (Cat. No. 01-2222-41, 01-2222-42) are stable for up to 3 days if the samples are stored in a fixative. If the samples are stored in FACS buffer, they are stable for 1 week.

Find additional tips, troubleshooting help, and resources within our Flow Cytometry Support Center.

I am performing flow cytometry. My compensation matrix has values over 100. That's not possible is it?

It is possible. But if you have a lot of values over 100, you probably need to look at the voltages that you are using for your data collection.

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

I’ve been told that I need to compensate my data for flow cytometry analysis. What does that mean?

By running single-color controls, it is possible to remove signal of one fluorophore that spills over into the collection channel for another fluorophore.

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

What kind of controls do I need for flow cytometry?

For compensation, you need to prepare a singly stained sample (or compensation beads) for each color parameter that you are using. In addition, we recommend that you use FMO (flow minus one) controls. These are controls in which you label cells or beads with every color in your panel, omitting one. Make one FMO control for each color. These controls are important for helping you properly set gates on your data.

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

Why should I worry about compensation in flow cytometry analysis?

In a perfect world, the fluorescence emission profile for each individual fluorophore would be a very intense, narrow peak, well separated from all other emission peaks. In reality, organic dyes and fluorescent proteins have broad emission peaks, and compensation must be employed (during or after data acquisition) to correctly assign fluorescence signal to each fluorophore. Compensation is important because it removes fluorescent signal from overlapping spectra so you know that the signal you see is only the signal from the fluorophore of interest.

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

How do I make compensation controls for antibodies?

All that you need for compensation controls is an unstained sample of your cells (or negative control beads) and samples with each of your fluorophores - one per tube. You want the single-stained controls to be as bright as you expect your brightest sample to be. Antibodies can be bound to beads instead of cells using our AbC Total Antibody Compensation Bead Kit (Cat. Nos. A10513 and A10497).

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

Can I use the OneComp and UltraComp eBeads microspheres with Super Bright-conjugated antibodies?

UltraComp eBeads microspheres (Cat. No. 01-2222) are recommended for use with Super Bright dyes.
Note: Super Bright Staining Buffer (Cat. No. SB-4400) is not compatible with UltraComp eBeads microspheres (Cat. No. 01-2222-41, 00-2222-42). If using UltraComp eBeads microspheres as a compensation tool, solely use Flow Cytometry Stain Buffer (Cat. No. 00-4222-26, 00-4222-57) for any antibody dilutions.

In some experiments, we have observed that compensation values for Super Bright 780- and Brilliant Violet 785- or Brilliant Violet 786-conjugated antibodies are higher in the violet 450/50 channel when using UltraComp eBeads microspheres as compared to single-color stained cells. In such circumstances, we would recommend setting compensation with cells. We have also observed this in some experiments using AbC Total Antibody Compensation beads, both with Super Bright 780 and Brilliant Violet 786. We have not tested Brilliant Violet 785 with the AbC beads.

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