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We reported a surface charge sensitive suspended nanoparticle crystal (S-NPC) as a promising nanofluidic-based sensing strategy. In the S-NPC, interspaces among nanoparticles construct a nanochannel network and provide nanofluidic electrokinetic properties. When electric double layers inside the interspaces are overlapped, conductance across the S-NPC will be directly related to the surface charge density of the nanoparticles, offering an approach to sense the surface charge. The enormous surface to volume ratio of the S-NPC yields very large electrical signals that facilities the sensing application. Having an easy and inexpensive fabrication approach and more importantly, being able to use off-the-shelf nanoparticles with different functional groups onside, the present S-NPC offers a potential of detecting various biological and chemical processes with surface charge coupling involved.