Mouse CD4/CD8 single Flourochrome Flow?

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beansta7's picture
Mouse CD4/CD8 single Flourochrome Flow?

In the mouse model, is it possible to observe both CD4 and CD8 populations by Flow Cytometry under a single laser/channel?

Li Li
Li Li's picture
Do you mean single laser or

Do you mean single laser or single channel? With signal laser, you may choose two color (channel) for double staining.
Li Li

tittletv's picture
I agree. We need more

I agree. We need more information about your question. The answer could be as simple as staining for CD3.

samm's picture
You can certainly observe

You can certainly observe both CD4 and CD8 with a single laser - look for the FITC and PE direct conjugates (e.g. from eBioscience, BD Pharmingen and others) if you want to use the standard 488nm Argon laser. You won't be able to see BOTH CD4 and CD8 on a single channel, but if it is only T cells you're looking for, just stain for CD3 or Thy1 (CD90?).

Fujiwara's picture
As said by samm, definitely

As said by samm, definitely you can observe both CD4/CD8 with a single laser. You could have a better definition of the population if you use different colors (e.g., FITC and PE). However, when you have available only antibodies with the same color, you could use both to detect the populations. The only thing, though, you have to titrate the antibodies in the manner you can separate in a histogram the brightest, dimmest and negative population. For example, you can use titrate (or use antibodies from different companies, sometimes they differ in the fluorochrome intensity) to have a histogram with three peaks, a bright CD4, a dimm CD8 and, of course, a negative population.
This is the principle that BD is using the CBA (cytometric beads array) to detect up to 6 cytokines using the same fluorochrome.

marcus muench
marcus muench's picture
A bit off topic but

A bit off topic but nonetheless,

Another method to get more info out of a limited number of channels is to use the same antibody in two colors, e.g. CD14-FITC+CD3-FITC+CD3-PE+CD19-PE. The trick is that this really only works well if you have two mutually exclusive populations such as CD14+ monocytes and CD19+ B-cells in blood in my example. Thus, all the double positives ar CD3+ T cells, which will have a distinct diagonal appearance. If the antibodies being added are the same clone or in any way can block each other, than it is important to add the two antibodies together, so that one doesn't block the other too much.