Generally, bacterial culture is performed manually and is subject to error. Here, we created a novel, wellordered and reliable system for dispensing bacteria microscopically by using paper and an ink-jet printer for controlled patterning. For paper to accommodate a culture medium, hydrophobic/hydrophilic patterns were incorporated onto the paper by immersing paper in a toluene solution of polystyrene and drying for complete hydrophobization, followed by etching discrete, small areas of hydrophilicity by ink-jet printing with toluene. Agar was hydrolyzed with sulfuric acid for appropriate viscosity and dispensed with an ink-jet printer. In a separate experiment, bacterial cells were sequentially printed on a medium and colonies were observed microscopically. The results of this experiment ensured the successful dispensing of bacteria using ink-jet printing. An almost constant number of particles per droplet were ejected using a polystyrene latex as a model of bacterial dispersion. Consequently, we expect this technology to be adapted for the development of a paper-based bioassay system.