Recently, in investigating the responses of T-cells from high responder mice that were primed with myoglobin (Mb) or with synthetic peptides containing antigenic site 5 and increasing in length stepwise by increments of two residues, we observed that T-cell recognition was highly dependent on conformation. In the present studies, tolerization experiments were carried out to further investigate this finding. Neonatal mice (BALB/cByJ) were either tolerized with Mb or with synthetic peptides of Mb containing antigenic site 5. Tolerization with Mb and subsequent immunization with Mb gave T-cells that did not proliferate in vitro to Mb or any of the peptides. T-cells from mice that were tolerized with a truncated peptide 139-153 (having deletions at Tyr-151 and Ala-144) and subsequently immunized with Mb proliferated in vitro to Mb and to peptides 132-153, 135-153 and 143-153. T-cells from mice that were tolerized with native Mb and subsequently immunized with peptides (which are unfolded in solution) did not proliferate in vitro to Mb, but responded well to the peptides. Conversely, tolerization with peptides had no effect on the recognition of, and the response to, native Mb by the T-cells, whereas the response to the peptides was completely removed. It was thus concluded that the recognition of protein antigens (or at least of Mb) by T-cells is (like the recognition by antibody) dependent on the conformation of the antigen.