When the museum was founded in 1974, its collection consisted of only 140 objects, donated by the National Museum of Damascus and the National Museum of Aleppo. The current collection (from 2011) consists of some 25,000 objects, including the majority of the clay tablets found at Mari. The museum also holds many objects found by international teams of archaeologists at sites in the Upper Khabur area, such as Tell Beydar, Tell Brak, Tell Leilan and Tell Mozan. The Euphrates Valley southeast of Deir ez-Zor is represented with objects from the Classical site of Dura-Europos, which was once a border city of the Roman Empire.
The exhibition halls cover an area of 1,600 square meters (17,000 sq ft) and are arranged around a courtyard. The construction of the new museum was a joint Syrian–German operation.
The collection is organized around five chronological themes: prehistory, ancient Syria (late fourth to first millennium BCE), the Classical period, the Islamic era and ethnography.