Milk fat content is an important parameter for milk quality assessment in the dairy industry. This study reports the development of a hand-held, highly sensitive fiber optic milk fat sensor using U-bent plastic optical fiber (POF) probes based on the refractive index (RI) of milk, an inherent physicochemical property of milk, which is significantly influenced by the milk fat content. The U-bent POF sensor probes facilitate dip type sensing and offer non-destructive assessment of milk quality in real-time. The sensor works on the principle of intensity modulation technique by exploiting evanescent wave absorbance and refractive losses. The fiber optic probes sense a change in RI of the medium as a result of attenuated total reflection by virtue of the evanescent wave interactions of light at the fiber core/cladding interface with the surrounding medium. A relative change in the refractive index property of milk as a result of variation in fat content and addition of water is investigated using U-bent POF probes for milk quality assessment. The sensor shows an optical absorbance sensitivity of 0.15 ΔA/Δ% fat for the variation of fat content in the milk. Besides, the response of the sensor to water addition in wide concentration range was studied. Furthermore, a low cost portable optoelectronic device is reported for instantaneous milk quality monitoring in farm fields. This sensor could be of huge potential in dairy industries, milk collection centers as well as end-user applications for real-time milk quality monitoring. © 2018 Elsevier Inc.