Flow Cytometry of Escherichia coli on Microfluidic Devices

Abstract

Flow cytometry of the bacterium Escherichia coli was demonstrated on a microfabricated fluidic device (microchip). The channels were coated with poly(dimethylacrylamide) to prevent cell adhesion, and the cells were transported electrophoretically by applying potentials to the fluid reservoirs. The cells were electrophoretically focused at the channel cross and detected by coincident light scattering and fluorescence. The E. coli were labeled with a membrane-permeable nucleic acid stain (Syto15), a membrane-impermeable nucleic acid stain (propidium iodide), or a fluorescein-labeled antibody and counted at rates from 30 to 85 Hz. The observed labeling efficiencies for the dyes and antibody were greater than 94%.

Two simple and rugged designs for creating microfluidic sheath flow

“…2,4 Additionally, cells and other sample components come into contact with the walls of the channel, which makes fouling a danger. [7][8][9][10][11] Ladisch and co-workers confined the sample on either side with air. 12 A variety of factors affected the size of the liquid stream, including the hydrostatic pressure and surface tension of the fluid.…”

“…A more robust approach, pioneered by Ramsey's group, 8,13,14 has been to confine the sample stream on either side with a particle-free aqueous solution. This design is simple and easily fabricated, leading several researchers to imitate it.…”

“…9,10,[15][16][17][18][19][20][21][22][23][24][25][26][27] Unfortunately, the sample still comes into contact with the top and the bottom of the channel, which necessitates the addition of a dynamic or covalent coating to mitigate the propensity for fouling. [8][9][10][11]25 Also, cells and particles can appear at any depth from the top to the bottom of the channel, so high numerical aperture optics still cannot be used.…”

Generation of highly uniform droplets using asymmetric microchannels fabricated on a single crystal silicon plate: Effect of emulsifier and oil types

“…Although the frequency of droplet generation can be as high as 12,000 Hz for water-in-oil droplets [4], the throughput in terms of volume flow rate of the disperse phase is very low because the droplets are formed from a single microchannel. These microfluidic devices are investigated for capillary electrophoresis [5], immunoassays [6], cellomics [7], proteomics [8], DNA analysis [9], etc.…”

Determining under‐ and oversampling of individual particle distributions in microfluidic electrophoresis with orthogonal laser‐induced fluorescence detection

“…Historically speaking, flow cytometry was the first technique to analyze individual subcellular particles [1], so it is not surprising that many of the first analyses of large populations of individual microparticles [2][3][4][5][6][7], cells [8,9], and subcellular particles [10] on microfluidic devices also used flow cytometry. Electrophoretic separation schemes have also been used since the first glass microfluidic devices were described by Manz et al [11] and Harrison et al [12,13] and they continue to be quite popular due to ease of application.…”