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. 2017 Nov 21;8:1591. doi: 10.3389/fimmu.2017.01591

Figure 2.

Figure 2

Simultaneous measurement of cytomegalovirus (CMV) pp65 antigen-specific CD8+ T cell responses presented by each of the six human leukocyte antigen (HLA) class I allotypes in an individual by enzyme-linked immunospot (ELISPOT). (A) Representative ELISPOT assay results. The frequency of CD8+ T cells secreting interferon-γ (IFN-γ) was measured using aAPCs (aAPC-HLA) and aAPC-pp65 (aAPC-pp65-HLA) transfected with individual HLA class I allotypes (HLA-A*02:07, -A*31:01, -B*07:02, -B*27:05, -C*01:02, and -C*07:02) of HD50. CD8+ T cells were seeded at 5 × 105 per well in one well and at 1 × 105 per well in two wells for each allotype and background groups. The background for aAPCs and aAPC-pp65 was defined as the frequency of IFN-γ secreting CD8+ T cells induced by each aAPC without HLA gene transfer. The frequency of IFN-γ secretion in CD8+ T cells, excluding background values, is presented. Finally, the frequency of pp65-specific T cells was determined as [(aAPC-pp65-HLA) − (aAPC-pp65)] − [(aAPC-HLA) − (aAPC)]. (B) Representative example showing non-specific responses in aAPCs expressing specific HLA allotypes (HLA-A*24:02 and -B*15:01) in HD30.