ABSTRACT Detection of and discrimination between different nanoparticles and biomolecules are vital steps in analytical, biochemical, and diagnostic biomedical procedures used in life sciences. Synthetic micro/nanopores in solid-state membranes form an emerging class of single-molecule detectors capable of detecting and probing the properties of particles and biomolecules with high throughput and resolution: The particles or biomolecules to be analyzed are added to an electrolyte solution in one of the two reservoirs of the detector system separated by a thin membrane containing a single micro/nanopore. An outer electric field induces an open-pore ionic current (Iopen) through the pore, dragging the particles with itself. Transient changes occur when a particle slightly smaller than the pore translocates through the pore. This electrical signal can be analyzed to derive information regarding to the particle or biomolecule size and even its morphology, concentration in the solution, and the affinity for the pore. Many detectors are based on self-assembled, naturally occurring protein pores in lipid bilayer membranes. Most solid-state pore-based detectors reported in literature use artificial pores in silicon nitride or silicon oxide membranes. Applying polymers as a membrane potentially offers advantages over the aforementioned types, including good electrical insulation, improved wettability thanks to higher hydrophilicity, and long-term stable yet low-cost and disposable devices. The present study aims at exploiting such advantages by developing the proof-of-concept for a single-material, all-polymer, nanopore detector allowing the continuous variation of target pore size in the range from micrometers to a few nanometers for best pore size adaption to the biomolecules to be investigated. The research comprises materials selection, system design, development of a fabrication and assembly sequence, device fabrication, and functional device testing. Poly (methyl methacrylate) (PMMA) was selected as it combines advantageous microfluidic properties know from competing materials, such as polyimide, polystyrene, polycarbonate, or polyethylene terephthalate, with outstanding micropatterning capabilities. The membrane thickness is set to be 1 µm, based on a compromise between robustness during fabrication and operation on one side, and electrochemical performance on the other. After spincoating the membrane onto a sacrificial wafer, pores with diameters of typically several hundred nanometers are patterned by electron beam lithography. In combination with thermal post processing leading to polymer reflow, diameters one order of magnitude smaller can be achieved. The present study focuses on 450 nm and 22 nm pores, respectively. Besides these pores fabricated in a top-down approach, self-assembled -hemolysin protein pores of 1.5 nm diameter are integrated in a combined top-down and bottom-up approach so that single digit, double digit, and triple digit nanometer pores are available. Systems integration is achieved by capillary-forced based release from the sacrificial substrate and the application of UV-initiated glue. Test sequences proved and qualified the device functionality: Electrical characterization was performed in aqueous KCl electrolyte solution. The devices exhibit a stable, time-independent ionic current. The current-voltage curves are linear and scale with the electrolyte concentration. System verification was performed using silica nanospheres of 100 nm and 150 nm diameter as known test particles. Translocation through a 450 nm pore induced current blockades for about 1 ms with an amplitude of 30 pA to 55 pA for 100 nm particles and in excess of 70 pA for 150 nm particles. This is in close agreement with results obtained by a mathematical model used in this study. Biomolecules relevant to many life science applications, double-stranded DNA (dsDNA) and bovine serum albumin (BSA) were subsequently analyzed to prove the device concept. Post-processed pores of 22 nm diameter were used at 600 mV driving voltage and 0.1 molar electrolyte in a slightly acidic regime of pH = 6. Typical current blockade amplitudes for complete translocations of dsDNA are Iblock = 22 pA for a translocation time of tD = 0.2 ms, and an almost threefold current blockade (Iblock = 60 pA) for the larger BSA molecules, respectively. The results demonstrate that the PMMA-based nanopores are sensitive enough to not only detect translocating biomolecules, but to also sense them by distinguishing between different biomolecules. The molecule-specific and distinct translocation signals through the pores using both, standardized silica nanoparticles and biomolecules of different dimensions, prove the concept of an all-PMMA electrophoretic flow detector with adjustable pore diameters. Devices with pore diameters covering three orders of magnitude in the nanometer range were successfully built, tested, and characterized. The results suggest such detectors are promising candidates for biomolecule detecting applications.