Targeted Bacterial Colony Isolation Using Light-Patterned Hydrogel Degradation

Take a coverslip with a photodegradable hydrogel layer that contains entrapped bacterial colonies, genetically modified to emit fluorescence.

Place the coverslip into a holder.

Add media to prevent sample dehydration.

Under a brightfield microscope, focus on a single colony of interest.

Using appropriate software, design a light pattern for cell extraction at the colony periphery to break down the structural bonds of the hydrogel and minimize UV exposure to the cells.

Adjust the UV light intensity and exposure duration.

Initiate UV light exposure to break the structural bonds in the hydrogel at the exposed areas that facilitate the release of the target colony.

Using a fluorescence filter, identify the colony.

Place a tubing tip at the spot, with the other end connected to a syringe.

Switch to brightfield mode and, using the syringe, aspirate the released colony.

Transfer the colony into a tube for further analysis.

Place the sample in a PDMS holder and add the defined media on top of the sample to prevent sample dehydration and provide a carrier solution for released cells. After bringing the calibration mirror in position, adjust the microscope focus to get a sharp image of the colonies wit