Cell agglomeration in the wells of a 24-well plate using acoustic streaming

Abstract

Cell agglomeration is essential both to the success of drug testing and to the development of tissue engineering. Here, a MHz-order acoustic wave is used to generate acoustic streaming in the wells of a 24-well plate to drive particle and cell agglomeration. Acoustic streaming is known to manipulate particles in microfluidic devices, and even provide concentration in sessile droplets, but concentration of particles or cells in individual wells has never been shown, principally due to the drag present along the periphery of the fluid in such a well. The agglomeration time for a range of particle sizes suggests that shear-induced migration plays an important role in the agglomeration process. Particles with a diameter of 45 μm agglomerated into a suspended pellet under exposure to 2.134 MHz acoustic waves at 1.5 W in 30 s. Additionally, BT-474 cells also agglomerated as adherent masses at the center bottom of the wells of tissue-culture treated 24-well plates. By switching to low cell binding 24-well plates, the BT-474 cells formed suspended agglomerations that appeared to be spheroids, fully fifteen times larger than any cell agglomerates without the acoustic streaming. In either case, the viability and proliferation of the cells were maintained despite acoustic irradiation and streaming. Intermittent excitation was effective in avoiding temperature excursions, consuming only 75 mW per well on average, presenting a convenient means to form fully three-dimensional cellular masses potentially useful for tissue, cancer, and drug research.

On‐Chip Generation of Vortical Flows for Microfluidic Centrifugation

“…This, in turn, drives the formation of a toroidal particle ring in the case of a particle‐laden droplet ( Figure ) . Additionally, it has also been shown that the placement of a piezoelectric element at an offset position beneath a chamber of a 24‐well microarray titer plate can be used to transmit the bulk vibration to the base of a well with the aid of a liquid couplant so as to generate a vortical flow in the chamber, although the complexity of the setup—in particular, the mounting of the element at an angle to the well and the electrical connections required to each element (one is required beneath each well)—makes realization difficult, and, as such, addressability of each well in the entire microarray plate was not demonstrated . A more elegant solution employing a hybrid combination of surface and bulk waves for on‐demand individual, sequential, and simultaneous addressability of all the wells in a 96‐well microarray plate will be discussed below …”

“…This was then demonstrated for generating microvortexing in the microwells—individually, sequentially or simultaneously—on‐demand for flexible addressability of the microwell titer plates as an attractive alternative to that in ref. ( Figure ) …”

“…• Discharge-driven [83][84][85][86][87] • Electrowetting [88] Ease of electrode fabrication and integration into microfluidic device if large applied potentials requiring bulky amplifiers are not required Electric field intensity can be tuned to control vortex characteristics and strength Chemical surface modification is an added complexity in channel fabrication; functionalization can wear off and needs to be reap- [89][90][91][92][93][94][95][96][97][98][99] -Posts/pillars/protrusions/ chamber [100][101][102][103][104][105] -Membranes [106] -Resonant cavities [107,108] -Microarray titer plate [109] -Plasmonic nanoparticles (photoacoustic effect) [110,111] • Direct contact with liquid [112][113][114][115][116][117] Surface vibration…”

“…[114,115] Additionally, it has also been shown that the placement of a piezoelectric element at an offset position beneath a chamber of a 24-well microarray titer plate can be used to transmit the bulk vibration to the base of a well with the aid of a liquid couplant so as to generate a vortical flow in the chamber, although the complexity of the setup-in particular, the mounting of the element at an angle to the well and the electrical connections required to each element (one is required beneath each well)-makes realization difficult, and, as such, addressability of each well in the entire microarray plate was not demonstrated. [109] A more elegant solution employing a hybrid combination of surface and bulk waves for on-demand individual, sequential, and simultaneous addressability of all the wells in a 96-well microarray plate will be discussed below. [139] Surface waves, in particular, surface acoustic waves (SAWs), which due to their higher frequencies (>10 MHz) and hence shorter wavelengths λ a ≪ h, are confined to the surface of the piezoelectric substrate of thickness h in contrast to the aforementioned bulk acoustic waves where λ a > h (λ a being the wavelength of the specific acoustic wave in the substrate), are an attractive alternative that have been shown to be particularly efficient in driving a wide range of microfluidic flows.…”

Microfluidic flow switching via localized acoustic streaming controlled by surface acoustic waves

“…The acoustic waves' attenuation produces a pressure gradient in the direction of wave propagation that generates a time-averaged body force (F B ) on the uid, thereby producing a streaming ow in the form of micro-vortices. Microvortices produced by SAWs have been widely used in microuidic platforms to mix uids, 26 sort particles, 27,28 manipulate cells, 29,30 drive SAW propulsion devices, 31,32 manipulate droplets, [33][34][35] and pump uids using acoustic ows.…”