PM2.5 filter samples (12-h and 24-h) were collected in urban Atlanta, GA, next to a freeway and 400 m away, as well as at a rural site, with particular focus on exploring on-road emissions, regional transport, and biogenic effects. Detailed speciation of PM2.5 carbonaceous aerosols was conducted by GC/ MS. Diurnal, seasonal, and spatial variations of PM2.5 organic composition were investigated. Primary organic compounds usually exhibit different attributes of day vs night while secondary organic tracers varied little. Much higher concentrations of automotive-related primary organic compounds are observed at the highway site, including n-alkanes, hopanes, steranes, and polycyclic aromatic hydrocarbons (PAHs). Season-specific on-road mobile source primary OC profiles were developed by using differences in organic species concentrations between the highway site and the nearby site. Calculated on-road source profiles differ from mobile source profiles measured in the lab. Significant seasonal differences are observed for 2-methyltetrols, cis-pinonic acid, and pinic acid, organic tracers of biogenic secondary organic aerosols (SOA). Little correlation is found between 2-methyltetrols and cis-pinonic or pinic acid, though cis-pinonic and pinic acids are strongly correlated.