Hi, I have immunised mice with a protein-based vaccine and will test the T cell response using IFN-γ ELISPot with overlapping peptides covering the antigen sequence.



	I plan to separate the CD8+ positive T-cells using magnetic cell separation prior to the ELISpot incubation. Do I need to add antigen presenting cells then? Since the rest of the splenocytes are excluded in the sorting. If so, do you have any recommended protocol? Is it possible to use murine primary dendritic cells?

Since the overlapping peptides will need processing my feeling is that antigen presenting cells will be needed. But I am not 100%. I will discuss the question with some other good immunologists. Edit: no I was wrong <img alt=":)" data-emoticon="true" height="20" src="//media.invisioncic.com/d244236/emoticons/default_smile.png" srcset="//media.invisioncic.com/d244236/emoticons/smile@2x.png 2x" title=":)" width="20">



	The purified T-cells should be able to present the peptides to each other without the need of APCs. 



	Adding murine APC is something I have not done previously in my career but my feeling is that most people work with in vitro matured DCs.

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Using APCs in ELISpot with CD8+ T cells