Manual multiple synthesis methods

Simultaneous multiple peptide synthesis enables the parallel synthesis of large numbers of peptides. The T-bag (tea-bag) method was developed along with other methods, e.g. pin synthesis, synthesis on paper plates, synthesis on parallel columns, and synthesis on cellulose, as technology to facilitate simultaneous multiple synthesis. Large numbers of peptides, peptidomimetics, and small organic molecules have been prepared using the T-bag method to address different research fields, such as conformational analysis, structure activity analysis, synthesis methodologies, and antibody-antigen interaction studies. Using the T-bag method, more than 150 peptides can be prepared in parallel in flexible amounts, with easily enough material for biological tests and analytical studies. The synthesis of peptides of length of up to 26 amino acid residues has been reported. Moreover, the T-bag technology is easy to apply in practice and requires very little special equipment. T-bags are prepared by containing solid phase resins within polypropylene mesh material. Polypropylene is rather chemically inert as well as fairly thermally stable (to 150°C), allowing a wide range of chemical reactions to be used for solid phase synthesis without affecting the bag material. Polystyrene cross-linked with 1% divinylbenzene, 100-200 mesh, is mainly used as the solid support, but other types of base resin can be used as well, e.g. TentaGel. The size of the resin beads must exceed the size of the pores of the polypropylene mesh material of the T-bags to avoid resin loss during synthesis. Syntheses are carried out manually, using semi automation, or within a multiple peptide synthesizer. The preparation of T-bags for solid phase synthesis, starting with 100 mg resin per bag, is described in Protocol 1. Synthesis using the T-bag method can be performed using either Boc or Fmoc synthetic strategies. For all manipulations, enough solvent should be used to cover the T-bags (about 3-4 ml per bag containing 100 mg of resin). To enable efficient washings and reactions, the reaction vessels (polyethylene bottles) should be shaken vigorously, preferably through the use of a reciprocating shaker.