This paper describes the development of a handheld device for long-term nanopore-based biosensing. Recently, membrane protein reconstituted in bilayer lipid membrane (BLM) has been applied for bio/chemical sensors because membrane protein intrinsically has a superior amplification mechanism. For mobile use of such devices, however, the bilayer requires mechanical robustness. Here, we propose a handheld BLM device aiming for a nanopore-based mobile sensor. BLM is formed at the tip of a glass capillary, which is contained in the solution trapped in a cup-like reservoir. This format achieved (i) formation of a sufficiently robust BLM for mobile use, (ii) solution exchange without rupturing BLM, and (iii) sustaining the small number of nanopores in the bilayer for accurate and long-term sensing. The handheld device successfully detected DNA translocations through the nanopores continuously for 75 minutes using 5 nanopores.