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J Immunol Methods. 2017 Feb;441:39-48. doi: 10.1016/j.jim.2016.11.011. Epub 2016 Nov 24.

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

Author information

1
Mabtech, Box 1233, SE-131 28, Nacka Strand, Sweden.
2
Swedish Orphan Biovitrum AB, SE-112 76 Stockholm, Sweden.
3
Departments of Virology and Immunology and Southwest National Primate Research Center, Texas Biomedical Research Institute, San Antonio, TX 78245-0549, USA.
4
Mabtech, Box 1233, SE-131 28, Nacka Strand, Sweden; Department of Immunology, Stockholm University, SE-106 91 Stockholm, Sweden. Electronic address: niklas@mabtech.com.

Abstract

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.

KEYWORDS:

Antibody; ELISA; ELISpot; Interferon-γ; Interleukin-2; Non-human primates

PMID:
27889562
PMCID:
PMC5563966
DOI:
10.1016/j.jim.2016.11.011
[Indexed for MEDLINE]
Free PMC Article

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