Thiopalmitoylation (i.e., the covalent attachment of palmitic acid via a thioester linkage to cysteine residues in the polypeptide backbone) is a common post-translational modification of proteins. Several proteins that have been identified as putative autoantigens in a variety of T-cell mediated autoimmune diseases are thiopalmitoylated, and thus, we have hypothesized that endogenous thiopalmitoylated peptides released during tissue breakdown may play a role in the development and chronicity of autoimmune diseases. To investigate this, we have studied the effect of thiopalmitoylation on the immunogenic and neuritogenic properties of P0, the major peripheral nervous system (PNS) myelin protein, which is thiopalmitoylated at cysteine 153, and described as a canditate autoantigen in Guillain-Barré syndrome (GBS), a human inflammatory demyelinating disease of the PNS. This paper describes the synthesis of palmitoylated peptide P0(180-199) and P0(152-171) by on-resin acylation using specific cysteine side-chain protecting groups: Mmt (labile in diluted acid) and StBu (labile in the presence of tributylphosphine). Our results show that the thiol protecting group had to be adjusted to the peptide sequence: Mmt was efficiently used for P0(180-199) thioacylation, but it was not suitable for thiopalmitoylation of P0(152-171) because of a premature deprotection of the Boc protecting group on the ε-NH2 Lys in the presence of 2% TFA, leading to dipalmitoylation. Palmitoylated P0(152-171) was successfully obtained by using StBu as the thiol protecting group. We could show by circular dichroism that palmitoylation has no influence on the structuration of the peptide in solution but palmitoylation increased the stability of the peptide in the presence of serum. Using EAN (experimental autoimmune neuritis), the rat model of GBS, we have compared the immunological properties of palm and non-palm P0 peptides and showed that thiopalmitoylation has indeed a great influence on their neuritogenic and immunogenic properties. This study provides further support for our hypothesis concerning the role of thiopalmitoylation in the development and chronicity of inflammatory demyelinating diseases and confirms that thiopalmitoylation of peptides may provide a simple means to increase MHC class II restricted responses.