Abstract: The objective of this research was to investigate the influence factors of testing contact angle of single fibers by using optical methods. The results indicated that the contact angles of fibers isolated by different methods were different, and the contact angle of fibers isolated in HNO3-KClO3 was different significantly with that isolated in NaClO and H2O2-HAc. With the same treatment, the fibers contact angle of different types was various, the smallest one was ramie fibers. Both temperature and relative humidity affected the contact angle of natural fibers, and humidity influenced more. Temperature affected contact angle of regenerated cellulose fibers scarcely, but humidity influenced significantly, however, temperature affected that of synthetic fibers little and humidity almost had no influence on it.