Mobile Protein Microboxes on Optical Fibers for Spatial Micromanipulation of Bacterial Microcolonies

Area of Science:

• Biotechnology and Bioengineering
• Microfluidics and Cell Culture
• Microscopy and Imaging

Background:

• Investigating intercellular interactions requires precise spatial control over cell populations.
• Previous methods using protein hydrogels for surface patterning had limitations.
• A new strategy is needed for active and reversible positioning of cells.

Purpose of the Study:

• To present a versatile strategy for actively and reversibly positioning cell populations.
• To enable three-dimensional, micrometer-scale manipulation of cells using microstructures.
• To create a platform for studying cellular behaviors influenced by spatial arrangement.

Main Methods:

• Fabrication of protein-based microstructures using two-photon polymerization.
• Printing permeable microboxes directly onto optical fiber tips for manipulation.
• Utilizing interlocking geometries, such as a 'Christmas tree' design, for stable docking with coverslip-mounted structures.

Main Results:

• Demonstrated controlled assembly and precise arrangement of distinct bacterial colonies (Pseudomonas aeruginosa PAO1 and PA14).
• Achieved stable, reproducible alignment and mechanical stability of docked microstructures.
• Successfully withdrew optical fibers without perturbing the assembled cell configurations.

Conclusions:

• The developed approach provides a generalizable platform for systematic investigation of cellular behaviors.
• Enables studies of chemical communication and interspecies signaling with submicrometer spatial precision.
• Offers new opportunities to understand how physical proximity influences intercellular interactions.