The invention discloses a device and a method for detecting the grain size of phytoplankton in seawater, wherein the device comprises a micro-fluidic chip bearing platform, a micro-fluidic chip, an excitation source component, a light detection component, a Coulter counting component, a data processing component, an underwater environment information collecting component, an automatic sampling component and a data transmitting and displaying component, wherein the micro-fluidic chip comprises a glass slide, a liquid storage hole A, a liquid storage hole B, a liquid storage hole C, a liquid storage hole D, a main channel, a detection channel, an impedance pulse sensing upstream detection channel and an impedance pulse sensing downstream detection channel. According to the invention, because the micro-fluidic chip acts as a detection micro device of grain size spectrum of phytoplankton, relevant photoelectric detection components and data processing components are small in volume, and the detection device internally contains a set of complete detection system including an automatic sampling component, a data transmitting and displaying component and the like, the device has the advantages of convenience in carrying, low cost, capability of detecting the grain size of underwater phytoplankton in the field in real time and the like.