Like most paper did, I used CD138 to detect mouse spleen plasma cells, but I find no difference between naive and stimulated mice and I can hardly find postive cells. What is the problem? Is there another way? Thanks!
One easy and definitive way of doing it is by a B cell ELISpot assay. With the ELISpot you will detect all Ab-secreting cells (depending on what Abs you coat the plates with) without the need for phenotyping. Here is a reference for a good method:
http://www.jimmunol.org/content/183/5/3373.long
If you must use FACS, you could do an intracellular staining instead. Normally you can see the Ab producing cells as they should be intracellular Ig high.
If you have access, the Blimp-GFP mouse (http://www.ncbi.nlm.nih.gov/pubmed/15492122) that one is also quite specific for plasma cells.
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You should also consider that you might be off with the timing of when you look at the B cell responses or that you actually don't have that many plasma cells coming from your immunization.
What are you immunizing with and at what time point do you look?
Thanks for your suggestions，Christopher. I immunized with OVA and detected 7d and 28d after immunization，I could detect high Ab titers in serum by ELISA
What adjuvant did you use and what dose of protein and route? I know in general OVA is a pretty poor antigen that give relatively low B cell responses (Beta-gal e.g. would give a stronger response).
It could be that much of the Ab response is an extrafollicular early responses. These are very short-lived and come up between day 3-5. Otherwise, I know for HIV-1 Env immunization a GC is normally observed between day 7-14 and then is waning off, so it could be that you miss most of it with the time points chosen.
If you can you should also do an Ag-specific stain. Likely your Ag-specific PCs are quite few even though you can detect Abs by ELISA.
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Anyway.. If possible for you, I would still recommend the ELISpot for a kinetics experiment to give you an idea what is the best time point to chose for the facs analysis.
Thanks, I will seriously consider your advices. In brief, I used aGalCer as adjuvant to activate NKT cells. NKT cells could provide efficient help to B cells, and I detected a GC response after 7d by intramuscular injection.
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