Figure 2.

Assessment of monoclonal anti-CD200 in relieving the inhibition of Th1 responses in CD200^hi AML patients with direct assessment of CD200R engagement on CD4⁺ memory T-cells. (a) Representative ELISPOT wells show the effect of anti-CD200 (+) or isotype control antibody (−) on IFNγ release following PPP stimulation for CD200^hi and CD200^lo AML patients. (b) Summary data illustrating IFNγ release in response to PPP stimulation as measured by ELISPOT for CD200^hi/lo AML patients +/− anti-CD200. (c) Representative flow cytometric plots illustrate CD200R expression on both CD4⁺ and CD8⁺ memory T-cells for AML patients (filled histograms represent immunoglobulin isotype-matched control and open histograms represent CD200R expression). Data represents mean±1 s.d., n=7. *P<0.05, analyzed by one-tailed paired t-test. (d) To directly assess CD200R engagement on CD4⁺ memory T-cells, the CD4⁺ memory T-cell clone; Belx2 were co-cultured with control or CD200⁺ K562 cells. The percentage of TNFα-producing Belx2 T-cell clones in co-culture assays in response to CD3/CD28 costimulation was measured by flow cytometry following intracellular staining. Summary data illustrates the frequency of TNFα-producing T-cells in CD200⁺ and CD200^- K562 co-cultures +/− anti-CD200. (e) Summary data illustrating MFI of the TNFα⁺ T-cell population in CD200⁺ and CD200^- K562 co-cultures +/− anti-CD200. Data represents mean±1 s.d., n=6. *P<0.05, analyzed by one-tailed paired t-test.