HTSU-IgG binding to unstimulated CD4⁺ T cells isolated from cord blood is restricted to the CD45RO⁺(memory) subset.

(A) Cord blood lymphocytes were enriched for CD4⁺ and CD8⁺ T cells as described in “Materials and methods,” and the binding assays performed. Panels i-iii, CD4⁺ T cells; panels iv-vi, CD8⁺ T cells. The level of HTSU-IgG binding was determined for unactivated cells (panels ii,v) and for cells activated for 5 days (panels iii,vi). Panels i and iv show the level of binding to negative control SUA-IgG. (B) Panels i-iii show freshly isolated cord blood lymphocytes enriched for CD4⁺ T cells that were sorted for CD45RO and CD45RA, and binding of HTSU-IgG to these cell populations from freshly isolated, unstimulated cord CD4⁺ T lymphocytes was determined. Panel i shows binding to CD45RA⁺ T cells from one donor, panel ii shows binding to CD45RO⁺ cells from the same donor as panel i. Panel iii shows binding to CD45RA⁺ T cells from a different donor. Panels iv and v show freshly isolated cord blood lymphocytes enriched for CD4⁺ T cells. Three-color flow cytometry was performed, using PE-labeled anti-CD62L antibody, PerCP-labeled anti-CD45RA antibody, and HTSU-IgG indirectly labeled with FITC as described in “Materials and methods.” Panel iv shows analysis performed by gating on the population positive for binding to the anti-CD45RA antibody. X-axis, binding of HTSU-IgG; y-axis, binding to anti-CD62L antibody. Panel v shows analysis performed by gating on the population positive for binding to the anti-CD62L antibody. X-axis, binding of HTSU-IgG; y-axis, binding to anti-CD45RA antibody.