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Is serum free medium ok in ELISpot?

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Hi Mabtech,

I just purchased the Human IL-2 ELISpot BASIC (ALP), product code: 3445-2A, and have some questions about the data sheet (see attached image).


1.    The medium in step B2, is it PBS or cell culture medium?
2.    “The same serum” in step B2 is what kind of serum?
3.    Does step B2 mean that the serum is added to the medium and the serum concentration is 10%?
4.    I use serum free medium for cell culture, is that ok?


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Hi D,

I have to begin by answering question number 4, which more or less answers the other questions as well: 
Yes, it is ok to use serum free medium, and in that case, there is no need to add any serum to the medium – “serum free medium” contains defined proteins as substitute for real serum. Here is a link to a paper by Janetzki et al (2010) showing very comparable ELISpot-results among all kinds of medium, serum and serum-substitutes https://www.ncbi.nlm.nih.gov/pubmed/19894047. The study was made using IFN-gamma ELISpot assays, but I would be very surprised if the findings can’t be extrapolated to all cytokine analytes, including IL-2.

So, at step B2, which is a step to block potential unspecific binding in the well, just add the serum free medium as it is to the well. The important thing here, and the point we want to make in the text of step B2, is that you use the same medium as used for the cell suspensions you are studying. This is because the cell culture continues in the ELISpot well during incubation time, and you don’t want to introduce anything new in the blocking step that might give rise to unspecific cell activation. 

Kind regards,

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