Inhalable particulate pollution has adverse effects on human health. Many studies have been conducted to investigate levels and risks of exposure to particulate pollution in public transit, but scant attention has been paid to variations in the pollution levels among different positions inside the cabin. Differences can be observed among different positions inside transit vehicles, like passenger density and activities, which may lead to differentiated exposure risks between different positions. Therefore, this study investigates the level of PM2.5 concentrations and the spatiotemporal variations among three different positions (front, middle, and rear) inside the cabin during bus travel. The three positions represent the three areas inside the bus cabin, i.e., front area (FA), middle area (MA), and rear area (RA). We developed a portable monitoring system for in-cabin measurements, which consists of a mobile device with the designated app, a PM2.5 collecting device, and a temperature-relative humidity monitor. Surveys were carried out on buses of a route in Shenzhen, China, which include inbound and outbound trips during both morning and evening peaks of two working days. Based on spatial and temporal information embedded in data collection devices, PM2.5 concentration data were spatially and temporally referenced. Comparative and descriptive statistical analyses were employed to examine the differences and variations in PM2.5 concentration among the areas inside the bus cabin. There are three major findings. First, for all of the surveyed trips, FA showed the highest and RA showed the lowest PM2.5 concentration. Second, concerning the average pollution level of route segments along the routes, the PM2.5 concentrations in the three cabin areas were higher during inbound trips than during outbound trips. Third, within route segments, PM2.5 concentrations in FA and MA increased rapidly after the bus stopped at stops, and then gradually decreased, while in RA they were relatively stable. In general, a higher level of passenger density increased in-cabin PM2.5 concentration. The findings of our study could provide insight into improving the bus in-cabin microenvironment, such as the seating arrangement for vulnerable passengers.