The synthesis of well-defined fragments of heparin (HP) and heparan sulfate (HS) is often hampered by difficult post-assembly manipulations on the fully elaborated oligosaccharides. In particular, the O- and N-sulfonation steps, usually performed separately, require long reaction times and often result in low yield due to problems associated with the incompletion of the reaction and the purification of highly sulfonated products, thus representing a bottle-neck in the synthesis. We report herein an effective microwave-assisted protocol for the simultaneous O,N-sulfonation of HP/HS-like saccharides. Complete O- and N-sulfonation were attained when using a SO3·NEt3 complex in a solvent mixture of NEt3/pyridine at 100 °C (MW heating) for 15 min, thus facilitating the purification process. Easy to implement, per-O,N-sulfonation of mono-, di-, tri- and tetra-saccharides with two to six reactive sites was performed effectively in short reaction times and excellent yields (>90%). Under smooth deprotection conditions, the resulting per-O,N-sulfonated saccharides were fully deprotected in high yields (>88%), providing saccharides pertinent to the synthesis of HP/HS-like fragments, including three tetrasaccharides relevant to the substrate of heparanase. Moreover, we developed a microwave-assisted protocol for the one-pot selective O-sulfonation/N-acetylation on disaccharide 7, which could be applied to synthesize the members of the GAG family bearing N-acetyl groups.