Bacterial Printing Press that Regenerates Its Ink:  Contact-Printing Bacteria Using Hydrogel Stamps

Abstract

This paper describes the use of micropatterned agarose stamps prepared by molding against PDMS masters to print patterns of bacteria on agar plates. Topographically patterned agarose stamps were inked with suspensions of bacteria; these stamps generated patterns of bacteria with features as small as 200 microm over areas as large as 50 cm2. Stamps with many small features (>200 microm) were used to study patterns of bacteria growing on media containing gradients of small molecules; stamps with larger features (>750 microm) were used to print different strains of bacteria simultaneously. The stamp transfers only a small percentage of cells that are on its surface to the agar at a time; it is thus possible to replica-pattern hundreds of times with a single inking. The use of soft stamps provides other useful functions. Stamps are easily customized to provide a range of patterns. When culture media is included in the agarose stamp, cells divide and thrive on the surface. The resulting "living stamp" regenerates its "ink" and can be used to pattern surfaces repetitively for a month. This method is rapid, reproducible, convenient, and can be used to control the pattern, spacing, and orientation between colonies of different bacteria.

Soft lithography for micro- and nanoscale patterning

“…1) 51 . Furthermore, µCP (with a stamp fabricated in gels rather than PDMS) can be used to directly print bacterial 52 or mammalian cells 53 .…”

Immunoimmobilization of Living Salmonella for Fundamental Studies and Biosensor Applications

“…Micro- (Xu et al 2007) and macro-contact printing (Weibel et al 2005) have been employed to transfer live bacteria onto the surface of a nutrient-rich matrix such as agarose or hydrogel. Bacterial microarrays have also been prepared by loading individual bacterial cells into microwells (2.5 µm wide, ~3 µm deep) at the distal end of an optical fiber bundle by centrifuge (Brogan and Walt 2005).…”

“…After the activated area is fully covered with bacteria, excess bacteria are released into the medium and become planktonic. This feature is not available for patterns prepared by embedding (Mbeunkui et al 2002;Polyak et al 2001;Premkumar et al 2002;Weibel et al 2005;Xu et al 2007) or physical entrapment (Kuang et al 2004). For these patterns, bacteria are held on the substrates or inside the microwells with weak forces, and the cells do not stay fixed to the patterned areas for long periods of time, causing the eventual disintegration of the pattern.…”

Building communities one bacterium at a time

“…These communities require techniques for controlling their spatial organization (e.g., geometry and distance) to achieve stability. One way to impose this organization is by using a microfluidic device (16,17), such as the system described by Kim et al (7), or other techniques for controlling the position of bacteria on surfaces (18,19).…”