Particle lithography has been extensively used as a robust and cost-effective method to produce large-area, close-packed arrays of nanometer scale features. Many technological applications, including biosensing, require instead non-close-packed patterns in order to avoid cross-talk between the features. We present a simple, scalable, single-step particle lithography process that employs colloidal self-assembly at liquid-liquid interfaces (SALI) to fabricate regular, open particle lithography masks, where the size of the features (40 to 500 nm) and their separation can be independently controlled between 3 and 10 particle diameters. Finally we show how the process can be practically employed to produce diverse biosensing structures.