FIG. 3.

3/46 supernatant reconstituted with fresh medium retains immunosuppressive activity. Purified CD4⁺ T cells were activated with plate-bound MAbs to CD3/CD28 or to CD3/CD46, and supernatants were harvested after 3 days. The supernatants were concentrated 10-fold using Amicon centrifugal filters with a 3-kDa molecular mass cutoff membrane and then reconstituted to the original volume using fresh serum-free medium. The reconstituted supernatants were then transferred to CFSE-labeled, BCG-infected PBMC from two donors. Absolute numbers of CD4 and CD8 responders (CFSE^lo IFN-γ⁺) were enumerated by flow cytometry after 7 days.