Native chemical ligation is carried out in aqueous solution and frequently gives near-quantitative yields of the desired ligation product. The challenge lies in the preparation of the necessary unprotected peptide-thioester building block. Peptide-thioesters are usually prepared by Boc chemistry SPPS; a thioester-containing peptide cannot be synthesized using a nucleophilic base, thus disfavoring Fmoc chemistry. Fmoc chemistry solid phase peptide synthesis techniques for generating peptide-thioesters are known; they make use of modifications of the Kenner 'safety catch' linker. In making peptide segments for use in native chemical ligation, protecting groups that release aldehydes or ketones should be avoided since these may cap the N-terminal cysteine. For the same reason, the use of acetone should be avoided, particularly prior to lyophilization and in washing glassware.