This article describes a method for generating and measuring bacterial growth using microdroplet culture chambers. The process involves encapsulating bacteria in microdroplets and monitoring their growth through optical density measurements.
Begin with a microdroplet culture chamber containing a microfluidic chip with pump-controlled connectors for fluid flow and waste removal.
Place a reagent bottle containing a bacterial suspension overlaid with oil inside the chamber.
Connect the bottle’s top tube to a pump port and the side tube to a chip connector. Close the chamber.
Set the growth curve parameters. Then, initiate the droplet generation.
As oil and bacterial suspension converge at the T-junction, the oil shear breaks the aqueous bacterial phase into uniform, bacteria-encapsulating microdroplets.
After droplet generation, remove the bottle, connect the chip connector to the pump port, and close the chamber.
As each droplet passes through the detection path, an optical fiber probe measures its optical density, representing the bacterial concentration.
Move the microdroplets back and forth to allow bacterial growth.
Over time, the optical fiber measures the droplet’s optical density, generating a growth curve.
Use a 10 milliliters sterile syringe to inject three to five milliliters of MMC oil from the syringe needle to the side tube into the reagent bottle. Then tilt and rotate the reagent bottle slowly to make the oil fully infiltrate the inner wall.
After injecting five milliliters of an initial bacteria solution into the bottle, fill the reagent bottle by injecting five to seven milliliters of the MMC oil. Pull out the independent quick connector A of the reagent bottle and insert the quick connector A into the bottle's quick connector B to complete the sample injection operation. Once done, open the door of the operation chamber to put the reagent bottle into the metal bath.
Pull out the C2 connector of the chip and the quick connector A of the reagent bottle. Connect the side tube connector of the reagent bottle to the C2 connector and the top tube connector to the O2 connector. Then, close the door of the operation chamber.
To choose the function of growth curve measurement, click on Growth Curve. In the parameter setting interface, input the Number as 15. Then, turn on the OD detection switch and set the Wavelength as 600 nanometers. Click on the Start tab to start droplet generation.
The process will take 15 minutes to complete.
When a pop-up window appears on the main interface prompting a message, open the door of the operation chamber to take out the reagent bottle and connect the C2 and O2 connectors. After closing the door, click the OK button in the pop-up window to automatically cultivate the droplets and detect the OD values.
As the growth curve reaches the stationary phase, click the Data Export button to export the OD data. Select the data save path and export the OD value recorded during the cultivation period in the dot CVS format. To plot the growth curve, use mapping software such as Excel and Origin 9.0.
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