IL-10
Description
IL-10 is a cytokine expressed mainly by monocytes with multiple, pleiotropic effects in immunoregulation and inflammation. It downregulates the expression of Th1 cytokines, MHC class II antigens, and co-stimulatory molecules on macrophages; it also enhances B cell survival, proliferation, and antibody production. IL-10 can block NF-κB activity, and is involved in the regulation of the JAK-STAT signaling pathway. IL-10 was initially reported to suppress cytokine secretion, antigen presentation and CD4+ T cell activation, but further investigation has shown that IL-10 predominantly inhibits lipopolysaccharide (LPS) and bacterial product mediated induction of the pro-inflammatory cytokines TNF-α, IL-1β, IL-12, and IFN-g secretion from Toll-Like Receptor (TLR) triggered myeloid lineage cells. IL-10 and related cytokines (IL-22) exert essential functions to maintain tissue homeostasis during infection and inflammation through restriction of excessive inflammatory responses, upregulation of innate immunity, and promotion of tissue repairing mechanisms. Their important functions in diseases are supported by data from many preclinical models, human genetic studies, and clinical interventions (Ouyang and O'Garra 2019). There are no available reagents that identify marmoset IL-10.
Alignment
Protein alignment for human, rhesus macaque and marmoset IL-10:
Protein alignment for marmoset, owl monkey, and squirrel monkey IL-10:
References
- Ouyang, W. and A. O'Garra (2019). "IL-10 Family Cytokines IL-10 and IL-22: from Basic Science to Clinical Translation." Immunity 50(4): 871-891.
Status
Antibody pairs that bind to different epitopes of marmoset (MAR) IL-10 have been identified. Clone MT-3515 is the monoclonal antibody (mAb) used for capture, while clone MT-32AD is used in a biotinylated form as detection mAb. These antibody pairs have been successfully tested in ELISA, ELISPOT, and LMX format. Capture antibody MT-3515 has been found to be functional both while immobilized by electrostatic forces to ELISA and ELISPOT plates or when covalently bound to polystyrene beads (Luminex assay).
The first validation assays for MT-3515/MT-32AD were performed using supernatants of NHP PBMC stimulated with a combination of lipopolysaccharide (LPS) and phytohemagglutinin (PHA). Table 1 shows an example of a LMX assay that includes reagents that detect CCL3 (MIP-1α), CCL4 (MIP-1β), IFN-γ, IL-10, IL-17A, and TNF-α simultaneously.
The values in the table are median fluorescence intensity (MFI) units, and show the capacity to recognize, in a simultaneous manner, the increase in concentration of six different cytokines in a single assay. The reagents against CCL-4, IFN-γ, IL-10, IL-17A, and TNF-α recognize these molecules across the human and selected Old World (OWM) and New World (NWM) monkey species. The reagents against CCL3 do not bind to NWM CCL3 with the same strength as they did for human and OWM CCL3. Additionally, stimulation of IL-17A production by PHA/LPS seems to be very intense for NWM cells, compared to human and OWM PBMC.
The second set of validation assays was performed with the samples generated in vivo after LPS challenge. The results are shown in Figure 1. Detection of plasma IL-10 after LPS IV challenge shows a transient 4-log increase in concentration, which returns to baseline levels at about 6 hours post-treatment. The pattern is similar for MAR, SQM, and OLM.
These assays complete the validation for the anti-IL-10 antibody pair MT-3515 (capture) and MT-32AD (detection), which can be used for the detection of soluble IL-10 in ELISA, ELISPOT, and LMX formats. These antibody pairs will be made available to investigators. Additional testing will be done to identify mAbs that can recognize IL-10 in intracellular cytokine staining assays.