Identification of Specific Cell Surface Markers on Immune Cells of Squirrel Monkeys (Saimiri sciureus)

Abstract

Nonhuman primates are an important experimental model for the development of targeted biological therapeutics because of their immunological closeness to humans. However, there are very few antibody reagents relevant for delineating the different immune cell subsets based on nonhuman primate antigens directly or with cross-reactivity to those in humans. Here, we report specific expression of HLA-DR, PD-1, and CD123 on different circulating immune cell subsets in the peripheral blood that included T cells (CD3+), T cells subsets (CD4+ and CD8+), B cells (CD20+), natural killer (NK) cells (CD3-CD16+), and natural killer T cells (CD3+CD16+) along with different monocyte subsets in squirrel monkey (Saimiri sciureus). We established cross-reactivity of commercial mouse antihuman monoclonal antibodies (mAbs), with these various immune cell surface markers. These findings should aid further future comprehensive understanding of the immune parameters and identification of new biomarkers to significantly improve SQM as a model for biomedical studies.

Figure 1

Enumeration of circulating levels of lymphocyte subsets. The different lymphocyte subsets (a) and monocyte subsets (b) as shown are identified by flow cytometry, and the frequencies are shown.

Figure 2

Expression on HLA-DR+ (a), PD-1+ (b), and CD123+ (c) on T cells, B cells, NK cells, and monocytes of squirrel monkey.

Figure 3

Expression of PD-1+, HLA-DR+, and CD123+ on subsets of T cells: CD3+ T cells (a), CD4+ T cells (b), and CD8+ T cells (c).

Figure 4

Expression of PD-1, HLA-DR, and CD123 on B cells: CD20+ B cells (a), NK cells (b), and NKT cells (c).

Figure 5

Expression of PD-1, HLA-DR, and CD123 on classical monocytes (a) CD14 and CD16, intermediate (b), and nonclassical monocytes (c).

Figure 6

Mean fluorescence intensity (MFI) of HLA-DR on T cells (a), HLA-DR on B cells (b), HLA-DR on monocytes (c), HLA-DR on NK cells (d); PD-1 on T cells (e), PD-1 on B cells (f), PD-1 on monocytes (g), PD-1 on NK cells (h); and CD123 on T cells (i), CD123 on B cells (j), CD123 on monocytes (k), CD123 on NK cells (l). Values are reported as not significant (ns),  ^∗p=0.032, or  ^∗∗p=0.0032.

Figure 7

Blocking of steady state PD-1 and HLA-DR expression on mitogen stimulation of squirrel monkey PBMC (a) and IFN-γ ELISPOT (b): PBMC (1 × 10⁵) were stimulated with Con A (2 µg/mL) for 72 hr in the presence and absence of PD-1 (5 µg/mL) or HLA-DR (Clone L243; 5 µg/mL) blocking antibodies. Stimulation index was calculated by division of absorbance (A540) of PBMC (+Con A) with absorbance of PBMC (−Con A) (a). PBMC (50,000) were stimulated with Con A (2 µg/mL) for 40 hr in the presence and absence of PD-1 (1 µg/mL) or HLA-DR (clone L243; 5 µg/mL) blocking antibodies for IFN-γ ELISPOT (b). In both experiments, an isotype control (IgG1 or IgG2a for PD-1 and HLA-DR, respectively) at the same concentration of PD-1 or HLA-DR was used as a control. Results from three animals are shown. Values are reported as not significant (ns),  ^∗p=0.032, or  ^∗∗p=0.0032.