In a relatively healthy patient population, circulating T cells will mostly have a resting or naive phenotype (CD45RO-, CD45RA+, CCR7+, CD62L+, CD27+, CD28+, IL7-Ra+) as opposed to the effector phenotype (CD45RO+, CD57+, PD1-, CD95+, CCR7-, CD62L-) and hopefully very few will have the exhausted effector phenotype (CD45RO+, CD57+, PD1+, CD95+, CD45RA-, CCR7-, CD62L-). Of course, a certain percentage of T cells will have the circulating memory T cell phenotype (central memory: CD45RO+, CCR7+ CD45RA- CD62L+; effector memory: CD45RO+, CCR7-, CD45RA-, CD62L-). [I have compiled these markers from a number of reviews and primary papers over the years.] Whether a particular medication causes T cells to be suppressed in their ability to respond to pathogens/antigens is an entirely different question and requires a more sophisticated approach than merely bulk phenotyping blood. Perhaps you might want to look at your medicated (and not) population just prior to and after immunization to see if T cells (and other effector cells like B cells) respond appropriately (you could use flow to measure expansion of appropriate populations). Or, you might like to remove the cells from the antigen naive but medicated patient and then ex vivo stimulate to see if they respond to antigen appropriately. In this case, you could use the CD107a (degranulation) assay or ELISPOT assays suggested above. It really depends on your hypothesis and what your lab/institution's infrastructure can support. Good luck!

To start with you can look at CD4, CD3 and CD8. CD3 are on all mature T cells, CD4+ are helper T cells and CD8+ are cytotoxic T cells. If you need more information on the different group, there is http://www.bdbiosciences.com/documents/tcell_brochure.pdf

You can look at cytokines as well.

Hi Yi Wen Qian. I know about the generalt CD markers. I was just wondering if there was a CD marker that was expressed when a t cell gets activated and vice versa. The CD3, 4 and 8 are only markers of a t-cell not related to the state of activation/suppression.

Cytokines could be a method to see that the different promotor of inflammation cytokines are downregulated during treatment...

Thanks :)

Sorry Lars, I miss read your question, I though you are asking about naive T cell activation. In your case cytokines is the best, you can use elisa blot, and there are kits like http://www.sabiosciences.com/ELISA_product/HTML/MEH-003A.html

Thanks :)

Hi Lars,

Do you want to measure directly ex vivo?

Or do you want to specifically stimulate the T-cells, and test whether they are still able to respond to the antigen?

If you want to measure all responses using flowcytometry, you could intracellularly stain for cytokines, such as IFN-gamma, IL-2 and TNF-alpha.

In addition, you can analyze CD154 (CD40L) or CD137 (4-1BB) upregulation.

As mentioned before, mostly several markers are combined to show T-cell activation.

From a clinician point of view, I recall that familiar phagocytic lymphoghagocytosis, a FACS test is used to see if CD8 pos cells can be activated. This is done by exposure to CD3 and labelling for the resulting expression of CD107a. In a normal individual you will see >15% of all CD8 cells now coexpressing CD107. Impaired T-cell activation, as seen in this disease, results in 1-5% only.

Check out this publication on the topic.

doi:10.1111/j.1365-2249.2010.04302.x

The strength of activation usually determines the level of expression of the various markers everyone proposed. While CD69 and CD25 expression requires weak activation (TCR stimulation alone), CD154 and HLA-DR usually requires much stronger activation (TCR and CD28 costimulation). Also, these markers peak at different times post-stimulation, CD69 peaks after only a few hours while CD154 peaks at 3-4 days after activation. You could also assess T cell proliferation by CFSE dilution or their functionality by intracellular cytokine (IFNgamma, IL-4) staining. CD8 T cell degranulation (as proposed by Matthias) is also a good idea. Suppressive markers such as CTLA-4 and PD1 could also be of interest to your studies. Good luck!

Ex vivo, use CD45RA and CCR7 for CD4s, and CD38 and HLA-DR for CD8s. In vitro, use cytokine production after PMA/ion stimulation for 4 hrs (with Brefeldin): TNF, IL2, IFNg and IL-17. Good luck.

Could I ask you if in your knowledge there's some important circulating CD markers of T cells? I know for example about the soluble CD25 but do you know if there are other types of soluble CD? Thanks

In a relatively healthy patient population, circulating T cells will mostly have a resting or naive phenotype (CD45RO-, CD45RA+, CCR7+, CD62L+, CD27+, CD28+, IL7-Ra+) as opposed to the effector phenotype (CD45RO+, CD57+, PD1-, CD95+, CCR7-, CD62L-) and hopefully very few will have the exhausted effector phenotype (CD45RO+, CD57+, PD1+, CD95+, CD45RA-, CCR7-, CD62L-). Of course, a certain percentage of T cells will have the circulating memory T cell phenotype (central memory: CD45RO+, CCR7+ CD45RA- CD62L+; effector memory: CD45RO+, CCR7-, CD45RA-, CD62L-). [I have compiled these markers from a number of reviews and primary papers over the years.] Whether a particular medication causes T cells to be suppressed in their ability to respond to pathogens/antigens is an entirely different question and requires a more sophisticated approach than merely bulk phenotyping blood. Perhaps you might want to look at your medicated (and not) population just prior to and after immunization to see if T cells (and other effector cells like B cells) respond appropriately (you could use flow to measure expansion of appropriate populations). Or, you might like to remove the cells from the antigen naive but medicated patient and then ex vivo stimulate to see if they respond to antigen appropriately. In this case, you could use the CD107a (degranulation) assay or ELISPOT assays suggested above. It really depends on your hypothesis and what your lab/institution's infrastructure can support. Good luck!

CD25, CD38, CD69 and HLA-DR are activation marker. you can use any one or combination of these depending on your experimental set up.

Is there any paper explaining differences between kinetics of CD40L and CD69?

Thanks!

CD69

In addition to these answers, I would suggest measure soluble CD30 in the serum of patients or cell culture supernatant. Moreover, using antibody against CD30 may be helpful.