Use of high frequency electrorotation to identify cytoplasmic changes in cells non-disruptively.

In this paper we demonstrate how the use of frequencies ranging from 50 kHz to 5 GHz in the analysis of cells by electrorotation can open the path to the identification of differences not detectable by conventional set-ups. Earlier works usually reported electrorotation devices operating below 20 MHz, limiting the response obtained to properties associated with the cell membrane. Those devices are thus unable to resolve the physiological properties in the cytoplasm. We used microwave-based technology to extend the frequency operation to 5 GHz. At high frequencies (from tens of MHz to GHz), the electromagnetic signal passes through the membrane and allows probing the cytoplasm. This enables several applications, such as cell classification, and viability analysis. Additionally, the use of conventional microfabrication techniques reduces the cost and complexity of analysis, compared to other non-invasive methods. We demonstrated the potential of this set-up by identifying two different populations of T-lymphocytes not distinguishable through visual assessment. We also assessed the effect of calcein on cell cytoplasmic properties and used it as a controlled experiment to demonstrate the possibility of this method to detect changes happening predominantly in the cytoplasm.

Efficient AC electrothermal flow (ACET) on-chip for enhanced immunoassays.

Biochemical reaction rates in microfluidic systems are known to be limited by the diffusional transport of reagents, leading often to lowered sensitivity and/or longer detection times in immunoassays. Several methods, including electrically powering electrodes to generate AC electrothermal flow (ACET) on-chip, have been adopted to enhance the mass transport of the reagents and improve microfluidic mixing. Here, we report a novel ACET electrode design concept for generating in-plane microfluidic mixing vortices that act over a large volume close to the reaction surface of interest. This is different from the traditional ACET parallel electrode design that provides rather local vertical mixing vortices directly above the electrodes. Both numerical simulation and experimental studies were performed to validate the new design. Moreover, numerical simulation was carried out to show the effects of experimental factors such as the reaction kinetics (association constant) and the reagent concentration on the ACET-enhanced surface-based assays. As a proof of concept, the new design for the ACET-enhanced immunoassays was used to improve the immunostaining signal of the HER2 (human epidermal growth factor receptor 2) cancer biomarker on breast cancer cells. Finally, the concept of scaling up the design has been validated by experiments (immunoassays on breast cancer cells for different ACET power and different assay times). In particular, we show that larger ACET in-plane designs can agitate and mix the fluid over large microfluidic volumes, which further enhances the immunoassay's output. We have achieved a 6-times enhancement in the assay signal with a 75% reduction in assay time.

Planar hydrodynamic traps and buried channels for bead and cell trapping and releasing.

We present a novel concept for the controlled trapping and releasing of beads and cells in a PDMS microfluidic channel without obstacles present around the particle or in the channel. The trapping principle relies on a two-level microfluidic configuration: a top main PDMS channel interconnected to a buried glass microchannel using round vias. As the fluidic resistances rule the way the liquid flows inside the channels, particles located in the streamlines passing inside the buried level are immobilized by the round via with a smaller diameter, leaving the object motionless in the upper PDMS channel. The particle is maintained by the difference of pressure established across its interface and acts as an infinite fluidic resistance, virtually cancelling the subsequent buried fluidic path. The pressure is controlled at the outlet of the buried path and three modes of operation of a trap are defined: idle, trapping and releasing. The pressure conditions for each mode are defined based on the hydraulic-electrical circuit equivalence. The trapping of polystyrene beads in a compact array of 522 parallel traps controlled by a single pressure was demonstrated with a trapping efficiency of 94%. Pressure conditions necessary to safely trap cells in holes of different diameters were determined and demonstrated in an array of 25 traps, establishing the design and operation rules for the use of planar hydrodynamic traps for biological assays.