Considering processing concerns regarding emerging food safety incidents and large-scale food testing, efficient and straightforward screening tools are urgently needed. The requirements for this technology include effective sample pretreatment, sensitive assay, and sample-to-answer solution that enable the detection of trace amounts of contaminants in complex food matrices. Here, we developed a chemiluminescence- and optical fiber-based fully automated immunosensing (COFFAI) system that conducts chemical contaminant food screening in a truly “sample-to-result” manner. Instead of utilizing laborious benchtop sample purification techniques followed by a time-consuming manual ELISA procedure, the user simply injects the sample of interest, and the whole procedure is automatically processed. We demonstrated that the utility of our COFFAI system can be used to detect trace amounts of chemical contamination in various milk samples within approximately 75 min. This method showed an ultrahigh sensitivity with a quinolone detection limit in the 0.022–0.065 µg/L range for skim milk, whole milk, and raw milk samples. The integration of pretreatment and assay into one automated system is an effective method for food testing. We believe that this platform provides a general method for efficient contaminant screening in foodstuffs. Display omitted •We designed a fully automated system that enables “sample-to-result” contaminant screening.•A novel immunoaffinity-based sample pretreatment method that does not require the use of an organic solvent was designed.•An optical fiber and charge-coupled device sensor was used for highly sensitive quantification.