"Bottom-up" method is a popular approach for the preparation of molybdenum disulfide quantum dots (MoS2 QDs) benefitting from less time consumption and no high-powered sonication required. But the relatively low fluorescent quantum yield of the obtained MoS2 QDs and the rare study about their in vivo behavior stimulate us to do more research in this area. In this paper, we proposed a "bottom-up" hydrothermal method to prepare MoS2 QDs with a quantum yield (QY) of 34.55% by optimizing a series of reaction conditions. The successful fluorescence imaging of tumor cells in vitro and in vivo as well as the systematic in vivo behavior study such as biocompatibility, biodistribution and metabolism route provided the good basis for their wider biomedical applications. (C) 2019 Elsevier Inc. All rights reserved.