Background: Prolonged standing during work causes a lower extremity pain and disorders. Patellofemoral pain syndrome (PFPS) is one of the common diagnoses of the knee pain. Although the etiology of PFPS is not completely understood, it is considered to be multifactorial. Objects: The purpose of this study was to investigate difference in strength of knee muscles, quadriceps:hamstring muscles strength ratio (Q:H ratio), asymmetry ratio of knee muscles strength and dorsiflexion range of motion (ROM) between standing workers with and without PFPS. Methods: Twenty-eight standing workers with PFPS and 26 age-, height-, and weightmatched standing workers without PFPS participated in this study. A tension sensor measured knee muscle strength, and motion sensor measured dorsiflexion ROM. The asymmetry ratio of knee muscles was calculated by a specific formula using the knee muscles strength of the dominant side and the sound side. An independent t-test was used to identify significant differences in the strength, ROM, Q:H ratio, and asymmetry ratio between the PFPS and normal groups. Results: The standing worker with PFPS have significantly lower dorsiflexion ROM (p < 0.000) and higher asymmetry ratio of the hamstring muscles strength (p < 0.000) compare to the standing worker without PFPS. No significant differences were seen in the strength of quadriceps muscle and hamstring muscles, Q:H ratio, and asymmetry ratio of quadriceps muscle strength. Conclusion: There was a significant difference in the asymmetry ratio of the isometric hamstring muscle strength. This finding suggests that the asymmetry ratio of isometric hamstring muscle strength may be more important than measuring only the hamstring muscle strength of the PFPS side. Furthermore, the results of this study showed a significant difference in dorsiflexion ROM between the standing industrial workers with and without PFPS. Dorsiflexion ROM and isometric hamstring muscle strength should be considered when evaluating the subjects with PFPS.