I wish to quantify cytokines in supernatant obtained lymphoproliferation assays but first I want to select which IL-12 should be tested.
Dear Javier:
Only the entire cytokine has biological activity, and that is the reason for which the IL-12p40 alone seems to have an antagonistic effect (because it blocks the interaction of IL-12p70 with its receptor).
Nevertheless, there are several cell types that can produce IL-12p35 but only DCs and macrophages produce IL-12p40 (and, at the end, the active IL-12).
In my case, I always measured IL-12p40 for this reason, and I believe it correlates fine with the active IL-12. But, as I said before, I have in mind and I conclude my results knowing that also IL-23 can be detected with my ELISA.
Finally, you should know that IL-12 is the major cytokine produced by DCs in vitro, so it is the best parameter to measure if you are thinking of producing and stimulating DCs from murine bone marrow cells or from human PBMCs.
Good luck!
IL-12 is a 70-kDa heterodimeric cytokine composed of two disulfide-bound subunits designated p35 (35 kDa) and p40 (40 kDa). IL-12p40 is thought to be a natural antagonist of IL-12p70, acting, at least in part, by competitive binding to the IL-12 receptor. Both in vitro and in vivo, the p40 subunit is secreted as a monomer or homodimer.
Dear Javier:
As Udaybhanu said, the cytokine IL-12 is composed by two different subunits: IL-12p35 (MW 35 kDa) + IL-12p40 (MW 40 kDa). So, the entire cytokine is cosidered IL-12p70 (because its total MW is around 70 kDa). But, you have to take into account that the subunit p40 is shared with other members of the IL-12 family: in this case, IL-23. So, if you are measuring IL-12p40 by ELISA you have to know that you are considering both cytokines (with similar but not identical biological fuctions).
In any case, make sure that you consider this while making your conclusions.
I hope that this information helps with your decision. Good luck!
Thanks for your answer! They are very helpful. Do you know which of the two induces production of INF-gamma in PBMCs? I want to do IL-12 ELISA but i don't know if IL-12p70 is the best of them.
Dear Javier:
Only the entire cytokine has biological activity, and that is the reason for which the IL-12p40 alone seems to have an antagonistic effect (because it blocks the interaction of IL-12p70 with its receptor).
Nevertheless, there are several cell types that can produce IL-12p35 but only DCs and macrophages produce IL-12p40 (and, at the end, the active IL-12).
In my case, I always measured IL-12p40 for this reason, and I believe it correlates fine with the active IL-12. But, as I said before, I have in mind and I conclude my results knowing that also IL-23 can be detected with my ELISA.
Finally, you should know that IL-12 is the major cytokine produced by DCs in vitro, so it is the best parameter to measure if you are thinking of producing and stimulating DCs from murine bone marrow cells or from human PBMCs.
Good luck!
Ok, thank you Matías, this information is very importan for my assays in vitro. Thanks again!
Dear Javier, il-12p40 is also believed to form homodimers p80 with own activity. You can check the work of A.M.Cooper in tuberculosis for more information,
IL 12 as INFgamma promote the Th1 activity, linked also to autoimmunity. In our research field, autism. increased levels of either IL 12 and INF gamma has been reported suggesting the stimulation of TH1 population and argue their involvement in the autoimmune process in ASD. Some studies published report also the associattion of mayor deterioro of core syntomps in autistic patients with higher levels of IL 12 reinforcing their role in the central autoimmunity in autism.
I have a dout, What could explain the not significant increase of INF gamma in the context of IL 12 significant increase in this patients?. Could be related to tanscription factor expression?.
HI, in case of evaluating immunomodulatory effects which of the IL-12 is most appropriate (IL-12p40 or IL-12p70 .
In our study on M.tuberculosis, using cytokine microarray, IL-12p40 significantly increases in smear positive cases as compare to smear negatives along with IP-10 and MIP-2 irrespective of other cytokines difference. This may indicate involvement of IL-12p40 in chemotactic action.
In other hand in our study IL-12p40 is increase in line with sTNFRII may implies the antagonist action of IL-12p40 through binding of beta sub-unite of IL-12.
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