Assessing Bacterial Motility Using a Microfluidic Device

Abstract

Source: Scheidweiler, D., et al. Combining Fluidic Devices with Microscopy and Flow Cytometry to Study Microbial Transport in Porous Media Across Spatial Scales. J. Vis. Exp. (2020).

This video demonstrates the use of a microfluidic device to quantify bacterial motility through a porous matrix composed of randomly arranged pillars. Fluorescently labeled bacteria are introduced into the device, imaged via microscopy as they move through the porous matrix into the observation channel, and analyzed to generate a breakthrough curve (BTC).

Protocol

1. Bacterial culture conditions

1. Working under a laminar flow hood, use 100 μL of a glycerol stock of green fluorescent protein (GFP)-tagged Pseudomonas putida KT2440 (1 × 10⁷ mL^-1, stored at -80 °C) to inoculate 5 mL of Luria-Bertani (LB) medium. Incubate at 30 °C while shaking at 250 rpm overnight.
2. The next day, resuspend 100 μL of the overnight culture in 5 mL LB medium and incubate under the same conditions for 5h (exponential phase). Sample a 1 mL aliquot into a 2 mL tube, allow to cool to room temperature (~15 min), and centrifuge (2300 x g for 5 min).
3. Remove the supernatant and a....

Results

Figure 1: Fluidic devices to study microbial transport in porous media (A) Illustration of a f.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Ethylenediaminetetraacetic acid (EDTA)	Sigma
Elastomer Sylgard 184	Dowsil	101697
Flow cytometer NovoCyte	Acea
Glucose	Sigma		https://www.makeblock.com/project/xy-plotter-robot-kit
Luria-Bertani (LB) broth	BD
Liquid dispenser, XY Plotter Robot Kit	makeblock
Microscope Axio Imager	Zeiss
Microscope AxioZoom v16	Zeiss
Microscope slides, 75 mm × 25 mm	Corning
Minipuls 3 peristaltic pump	Gilson
Plasma bonder Corona SB	BlackHole Lab
Potassium phosphate	Sigma
Syringe pump New Era NE 4000	New Era
Syto 13 Green Fluorescent Nucleic Acid Stain	Molecular Probes, Invitrogen
Tygon tubing	Ismatec
WF31SA universal milling machine	Mikron