Conventional thin film nanocomposite (TFNC) membranes are based on porous membranes produced by the phase inversion method. The top barrier layer in these membranes has smaller pores with a torturous pathway and together with fairly low porosity for the supporting layer, resulting in a relatively low flux. In this study, a high flux ultra-filtration cellulose nanofiber-based (CN) membrane which consists of a three-tier composite structure, consisting of a TEMPO-oxidized cellulose nanofiber top layer, an electrospun poly(acrylonitrile) (PAN) scaffold and a non-woven polyethylene terephthalate (PET) support was used as a substrate for the thin film nanocomposite membrane preparation. The properties of the cellulose nanofiber-based membrane were fully characterized. The barrier layer of this membrane was prepared by interfacial polymerization (IP) of m-phenylenediamine (MPD) and piperazine (PIP) with trimesoyl chloride (TMC) on top of the CN membrane. The interfacial polymerization between MPD and TMC was investigated by studying the effects of the MPD concentration change, reaction time and curing temperature. Besides, the addition of PIP into the aqueous phase greatly improves the permeate flux without sacrificing the rejection ratio. When the MPD and PIP concentration were 1.5% and 0.5%, respectively, the thin film nanocomposite membrane with CN substrate exhibited a rejection of 94.6% and a permeate flux of 30.5 L/m2h, about 2 times higher than that of only 2% MPD concentration. Such thin film nanocomposite membrane has about 30% lower permeate flux than the commercial membrane, Dow FilmtecXLE-440 with a comparable rejection ratio (~95%) due to the thicker barrier layer produced by a manual preparation process. In addition, the A and B values of this TFNC membrane were 4.3 ?? 0.1 L/(m2*h*bar) and 1.54 L/m2h, compared with the commercial membrane, Dow filmtec XLE-440, 6.9 L/m2h*bar and 3.06 L/m2h, respectively. The filtration performance of this thin film nanocomposite membrane under various applied pressures (100~800 psi) was also studied.