The lithium anode could greatly increase the energy density of the cell, but inevitable lithium dendrites hinder its application. A powerful coating layer can effectively suppress the growth of dendrite by promoting a fast and uniform Li+ flux. Here, a polyolefin separator coated with a layer of nano cellulose fiber (NCF) and graphene oxide (GO) is designed to restrain the growth of Li-dendrite and accommodate thermal shrinkage resistance. The results of electrochemical impedance spectroscopy showed that the NCF/GO coating with a mass ratio of 3:5 showed the highest value of ionic conductivity. The Li/separator/Li symmetrical battery with NCF/GO coated separator can work for 65 h during continuous charging before the voltage dropping down to zero, which is about 40% longer than the battery with bare polyolefin separator. In addition, the NCF/GO coating layer can also increase the retention capacity ratio of a cell by more than 10% in the 100 charge/discharge cycles. In this paper, the as-prepared NCF/GO coating separator provides a reference for designing a long-life and safety battery with Li-anode by effectively inhibiting the lithium dendrite.