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Web Date: October 3, 2012

Microfluidic Device Synchronizes Bacteria

Cellular Analysis: Automated method produces bacterial populations in a single stage of the life cycle
Department: Science & Technology | Collection: Life Sciences
News Channels: Analytical SCENE, Biological SCENE
Keywords: bacterial synchronization, microfluidics, Caulobacter crescentus, cell cycle
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Cycle Of Life
Flagellum-bearing cells of the bacterium species Caulobacter crescentus get ready to pinch off from their immobile parent cells.
Credit: Courtesy of Yves Brun
micrograph of Caulobacter crescentus cells
 
Cycle Of Life
Flagellum-bearing cells of the bacterium species Caulobacter crescentus get ready to pinch off from their immobile parent cells.
Credit: Courtesy of Yves Brun

Left to their own devices, bacterial populations contain a mix of young, middle-aged, and old cells. But scientists studying events that occur at different parts of the microbes’ cell cycles learn more about each phase by studying populations all at the same stage. Researchers have now developed a microfluidic device that automatically syncs bacteria to the same stage for on-chip analysis (Anal. Chem., DOI: 10.1021/ac301565g).

The life cycle of the freshwater bacterium Caulobacter crescentus contains two morphologically different cell types: a sticky immobile stalked cell that divides to release a mobile flagellated cell called a “swarmer.” Its two cell types make the bacterium “a good model system” to study bacterial cell cycles, says Stephen Jacobson of Indiana University, Bloomington. For instance, the swarmer cells adhere to surfaces where they become stalked cells and form a biofilm. But, Jacobson says, scientists don’t understand how the cells become stuck in the first place. “To study that process, it’s best to synchronize the cells,” he says.

The current method for synchronizing C. crescentus is called the plate-release technique, says Jacobson. Researchers grow the bacteria on a petri plate and then add buffer to pick up newborn swarmers. By collecting the buffer containing synchronized swarmers, the researchers leave older stalked cells stuck to the plate. The drawbacks of plate release, according to Jacobson, are that the process is labor intensive and that scientists can collect synchronized populations only for a day before the plate becomes overgrown. When that happens, Jacobson says, the buffer starts picking up stalked cells as well as swarmers, so the resulting sample contains a mix of cell types.

Jacobson and colleagues aimed to automate the production, collection, and analysis of synchronized cells using a microfluidic device containing a series of chambers. They started by pumping unsynchronized cells from an input chamber into a central serpentine chamber with a large surface area for stalked cells to latch onto. As the biofilm grew, the researchers prevented overgrowth by flushing the chamber every 15 minutes with media the microbes need for growing. After a day, they started collecting synchronized swarmers by pumping media into the chamber, allowing it to gather newborn cells for five minutes, and then sending the fluid into an analysis chamber.

Because the C. crescentus cells expressed green fluorescent protein, the researchers could follow the bacteria’s movement in the analysis chamber on the chip and thereby check their synchronization using a fluorescence microscope. They found that 89% of the cells captured by the fluid were swimming—synchronized swarmers. The team confirmed their results with an off-chip analysis using flow cytometry. This percentage is similar to that obtained by plate release, which was 87%. Unlike in plate release, the microfluidic device kept producing synchronized populations of cells for four days, after which the biofilm became overgrown.

Compared to plate release, Shuichi Takayama of the University of Michigan, Ann Arbor, says, “They can sample more frequently, use less cells, and less material.” As a result, scientists can study synchronized cells more easily and cheaply. Jacobson is planning experiments that probe the formation of bacterial biofilms. He says the device should work with any cell type that, in one phase of its life, sticks to the device’s walls naturally--or that can be made to adhere.

 
Chemical & Engineering News
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