The city may have been the home of the Physician Luke who traveled with Paul on occasion. As a result, Luke may have taken special interest in his description of the city as the "capitol of first district of Macedonian Rome" (Acts. 16:12) a reference to the historical division of Macedonia earlier in the Roman period. In addition to its historical importance, Philippi was located along the important "Via Egnatia", the Roman road from Asia Minor that traversed the Balkan Peninsula toward the Adriatic Sea to ports with direct passage to Italy. In a sense, Philippi was the great roadway garrison station for the "eastern gate" from Europe to the Persian cities. By Roman times, the city had two types of citizens: Italians commissioned to live here and "political proselytes" like Paul and Silas, who were brought into the Roman citizenry by legislation of Rome.

Archaeologists have carefully constructed a working model of the occupation of the site. The earliest periods (Neolithic to Early Bronze) yield evidence of a settlement referred to as the "Dikli-tach", a group that used the flood plain for agriculture eventually gave way to a Thracian culture settlement. The city was founded as early as 700 BCE, and the site was well populated by both Thasos and the Thracian peoples during the Classical Period. The Classical period name of the site was CRENIDES (fountains), possibly because of a large ornament in the city square or the vast amount of surface water.

As King Philip II of Macedon took complete control of the region (365 BCE, control after 358/7 BCE) as a border garrison fort against Thrace, he swept in to dominate the nearby gold mines at Mt. Pangaeus. The gold was used to finance the build up of Philip's (and later Alexander's) army. With the rise of Rome, King Perseus (last of the Macedonian Kings) was routed from the Macedonian throne and ceded the area to Roman control. The Romans initially divided the area into four districts, later reorganizing Macedonia by 148 BCE as a single province.