Inhibitor formation in patients with haemophilia receiving factor VIII (FVIII) concentrate is a common problem requiring tolerance induction therapy. Immune tolerance is dependent on defective T cell/antigen-presenting cell (APC) interactions and inhibitor antibody formation is associated with effective T-cell/B-cell interaction. We studied the expression of the cell-surface molecules involved with these interactions using multiparameter flow cytometry and a whole blood stimulation assay-phytohaemaglutinin (PHA) to activate T cells and Escherichia coli lipopolysaccharide (LPS) to activate monocytes and B cells. Up-regulation of T-cell co-stimulatory receptors CD11a, CD40 ligand (CD40L) and CTLA4 were inhibited in a dose-dependent manner by plasma-derived (pd)FVIII, but CD28 was unchanged. Up-regulation of monocyte and B-cell co-stimulatory ligands CD4O, B7-1 (CD80) and B7-2 (CD86) were also inhibited in a dose-dependent manner by pdFVIII, but LFA-3 (CD58) was unchanged. The combined inhibitory effect of prednisolone, an immunosuppressive agent used in several tolerance induction protocols, with pdFVIII on co-stimulatory molecules, was additive. There was no significant alteration in T-cell/APC adhesion or co-stimulatory molecules noted in the presence of recombinant (rh)FVIII concentrate. The inhibitory effect of pdFVIII on molecules involved in interaction between T cells and APCs may result in immune tolerance in recipients of pdFVIII concentrate. The inhibitory effect of pdFVIII on CD40/CD40L up-regulation may result in defective antibody formation. We now provide evidence that the use of pdFVIII, through interfering with APC/T-cell interactions, may be more appropriate than rhFVIII for tolerance induction.