Systematic evaluation of monoclonal antibodies and immunoassays for the detection of Interferon-γ and Interleukin-2 in old and new world non-human primates

•Panels of mAbs to human IFN-γ and IL-2 were assessed for cross-reactivity with NHP.•NHP included both Old and New World species.•Pan-reactive mAbs recognizing cytokines from all NHP species were identified.•Capture and detection mAb pairs reactive with all NHP species were developed.•Functionality of mAbs was verified in ELISA, ELISpot, FluoroSpot and flow cytometry.

Non-human primates (NHP) provide important animal models for studies on immune responses to infections and vaccines. When assessing cellular immunity in NHP, cytokines are almost exclusively analyzed utilizing cross-reactive anti-human antibodies. The functionality of antibodies has to be empirically established for each assay/application as well as NHP species.A rational approach was employed to identify monoclonal antibodies (mAb) cross-reactive with many NHP species. Panels of new and established mAbs against human Interferon (IFN)-γ and Interleukin (IL)-2 were assessed for reactivity with eukaryotically expressed recombinant IFN-γ and IL-2, respectively, from Old (rhesus, cynomolgus and pigtail macaques, African green monkey, sooty mangabey and baboon) and New World NHP (Ma's night monkey, squirrel monkey and common marmoset). Pan-reactive mAbs, recognizing cytokines from all NHP species, were further analyzed in capture assays and flow cytometry with NHP peripheral blood mononuclear cells (PBMC).Pan-reactive mAb pairs for IFN-γ well as IL-2 were identified and used in ELISA to measure IFN-γ and IL-2, respectively, in Old and New World NHP PBMC supernatants. The same mAb pairs displayed high functionality in ELISpot and FluoroSpot for the measurement of antigen-specific IFN-γ and IL-2 responses using cynomolgus PBMC. Functionality of pan-reactive mAbs in flow cytometry was also verified with cynomolgus PBMC.The development of well-defined immunoassays functional with a panel of NHP species facilitates improved analyses of cellular immunity and enables inclusion in multiplex cytokine assays intended for a variety of NHP.