In this work, buckypapers (BPs) were synthesised by different types of non-ionic surfactants. The prepared BPs were characterised by different methods. Using of non-ionic and also non-toxic surfactant, Tween 20, made the surface possible to immobilise the biological molecules on the surface for measuring the electrochemical properties changes. Moreover, this surface showed better electrochemical behaviour, higher conductivity, and better charge transfer kinetics than BP which was synthesised by conventional Triton X100. To our knowledge, this is the first research on the effect of surfactant type on the BP synthesis and characteristics.