Characterization of small microfluidic valves for studies of mechanical properties of bacteria

Abstract

Lab-on-a-chip platforms present many new opportunities to study bacterial cells and cellular assemblies. Here, a new platform is described that allows application of uniaxial stress to individual bacterial cells while observing the cell and its subcellular assemblies using a high resolution optical microscope. The microfluidic chip consists of arrays of miniature pressure actuated valves. By placing a bacterium under one of such valves and partially closing the valve by externally applied pressure, the cell can be deformed. Although large pressure actuated valves used in integrated fluidic circuits have been extensively studied previously, here those microfluidic valves are downsized and flow channels with rectangular cross-sections are used to maintain the bacteria in contact with cell culture medium during the experiments. The closure of these valves has not been characterized before. First, these valves are modeled using finite element analysis, and then the modeling results are compared to the actual closing profiles of the valves, which is determined from absorption measurements. The measurements and modeling show with good agreement that the deflection of valves is a linear function of externally applied pressure and the deflection scales proportionally to the width of the flow channel. In addition to characterizing the valve, the report also demonstrates at a proof-of-principle level that the device can be used to deform a bacterial cell at considerable magnitude. The largest deformations are found in 5 μm wide channels where the bacterial width and length increase by 1.6 and 1.25 times, respectively. Narrower and broader channels are less optimal for these studies. The platform presents a promising approach to probe, in a quantitative and systematic way, the mechanical properties of not only bacterial cells but possibly also yeast and other single-celled organisms.

Mechanical slowing-down of cytoplasmic diffusion allows in vivo counting of proteins in individual cells

“…1d ) instead of syringe pumps to allow for easy streamlining and fast response times. Although MACS essentially exploits valve actuation to immobilize cells between a glass coverslip and a PDMS membrane similar to what was described earlier 23 24 , simply collapsing the valve (that is, going directly from open to closed state) yields extremely poor trapping efficiency due to the rapid displacement of liquid. Instead, MACS relies on three distinct valve states, achieved by controlling both the pressure of the valve ( P valve ) and the pressure driving the flow ( P flow ) of the cell suspension.…”

Analysis of Factors Limiting Bacterial Growth in PDMS Mother Machine Devices

“…In deformed regions of the channel, the cells grow in multiple rows or were tilted relative to the direction of the channel. Based on our earlier studies (Yang et al, 2015 ), stress analysis of such channels has shown that pressures in the 0.2 MPa range are needed to widen channels at their midplane by few hundred nanometers. Similar or even larger pressures must have been present in the broadened regions of the channel.…”

“…The channels in PDMS elastomers are created using 4″ silicon (Si) wafer molds. The fabrication of the Si molds follows the process described earlier (Yang et al, 2015 ). Briefly, the patterns of dead-end channels are defined by e-beam lithography using a JEOL JBX-9300FS electron beam lithography system (JEOL, Japan) with ZEP520A, a positive tone e-beam resist (ZEON Chemical, Japan).…”

“…In the latter case bacteria grow packed side-by-side and quantitative analysis of individual cells is more complicated. In addition to providing a steady growth environment, microfluidics can also be used to administer different chemical (Baltekin et al, 2017 ; Kaiser et al, 2018 ) and physical stimuli (Yang et al, 2015 ) to the cells in situ while they are imaged under the microscope.…”