Noncontact Multiphysics Probe for Spatiotemporal Resolved Single-Cell Manipulation and Analyses.

Heterogeneity and spatial arrangement of individual cells within tissues are critical to the identity of the host multicellular organism. While current single-cell techniques are capable of resolving heterogeneity, they mostly rely on extracting target cells from their physiological environment and hence lose the spatiotemporal resolution required for understanding cellular networks. Here, a multifunctional noncontact scanning probe that can precisely perform multiple manipulation procedures on living single-cells, while within their physiological tissue environment, is demonstrated. The noncontact multiphysics probe (NMP) consists of fluidic apertures and "hump" shaped electrodes that simultaneously confine reagents and electric signals with a single-cell resolution. The NMP's unique electropermealization-based approach in transferring macromolecules through the cell membrane is presented. The technology's adjustable spatial ability is demonstrated by transfecting adjacent single-cells with different DNA plasmid vectors. The NMP technology also opens the door for controllable cytoplasm extraction from living single-cells. This powerful application is demonstrated by executing multiple time point biopsies on adherent cells without affecting the integrity of the extracted macromolecules or the viability of cells. Furthermore, the NMP's function as an electro-thermal based microfluidic whole-cell tweezer is reported. This work offers a multifunctional tool with unprecedented probing features for spatiotemporal single-cell analysis within tissue samples.

Ventilation-Based Strategy to Manage Intraoperative Aerosol Viral Transmission in the Era of SARS-CoV-2.

In operating theaters, ventilation systems are designed to protect the patient from airborne contamination for minimizing risks of surgical site infections (SSIs). Ventilation systems often produce an airflow pattern that continuously pushes air out of the area surrounding the operating table, and hence reduces the resident time of airborne pathogen-carrying particles at the patient's location. As a result, patient-released airborne particles due to the use of powered tools, such as surgical smoke and insufflated CO2, typically circulate within the room. This circulation exposes the surgical team to airborne infection-especially when operating on a patient with infectious diseases, including COVID-19. This study examined the flow pattern of functional ventilation configurations in view of developing ventilation-based strategies to protect both the patient and the surgical team from aerosolized infections. A favorable design that minimized particle circulation was deduced using experimentally validated numerical models. The parameters adapted to quantify circulation of airborne particles were particles' half-life and elevation. The results show that the footprint of the outlet ducts and resulting flow pattern are important parameters for minimizing particle circulation. Overall, this study presents a modular framework for optimizing the ventilation systems that permits a switch in operation configuration to suit different operating procedures.

Airplug-mediated isolation and centralization of single T cells in rectangular microwells for biosensing.

Sorting cells in a single cell per microwell format is of great interest to basic biology studies, biotherapeutics, and biosensing including cell phenotyping. For instance, isolation of individual immune T cells in rectangular microwells has been shown to empower the multiplex cytokine profiling at the single cell level for therapeutics applications. The present study, however, shows that there is an existing bias in temporal cytokine sensing that originates from random "unpredicted" positions of loaded cells within the rectangular microwells. To eliminate this bias, the isolated cells need to be well-aligned with each other and relative to the sensing elements. Hence, an approach that utilizes the in situ formation and release of airplugs to localize cells towards the center of the rectangular microwells is reported. The chip includes 2250 microwells (each 500 × 50 × 20 µm3) arranged in 9 rows. Results showed 20% efficiency in trapping single T cells per microwells, where cells are localized within ±3% of the center of microwells. The developed platform could provide real-time dynamic and unbiased multiplex cytokine detection from single T cells for phenotyping and biotherapeutics studies.

Microelectrofluidic probe for sequential cell separation and patterning.

Cell separation and patterning are of interest to several biological and medical applications including rare cell isolation and co-culture models. Numerous microfluidic devices have been used for cell separation and patterning, however, the typical closed channel configuration comes with challenges and limitations. Here, we report a dielectrophoresis (DEP) enabled microelectrofluidic probe (MeFP) for sequentially separating and patterning of mammalian cells in an open microfluidic system. The MeFP is a microfluidic probe with injection and aspiration apertures, integrated with an array of micro-hump electrodes on its tip. Aligning the MeFP parallel, and in close proximity, to a conductive substrate forms a vertical pin-plate electrode configuration that allows for an integration of DEP forces within the hydrodynamic flow confinement. Upon confining a heterogeneous cell suspension in the gap between the MeFP and the substrate, target cells are selectively captured on the micro-hump electrodes using positive DEP forces, and then deposited on the substrate in defined patterns. Characterization of the MeFP showed an increase in cell-capture efficiency when the MeFP is of a higher microfluidic multipole configuration. Separation of cancer cells from T lymphocytes was demonstrated with capture purity as high as 89.6%. Deposited patterns of isolated cells match the numerically calculated particle trajectories of the evaluated microfluidic multipoles configurations. By adjusting the flow configuration of the MeFP, we show that the patterned co-culture of two different cell types can be dynamically controlled for homotypic and heterotypic cell interaction studies. This work presents a multifunctional microfluidic tool that bio-fabricates selective multicellular patterns directly on an open substrate without the need for confined conduits.

An integrated adipose-tissue-on-chip nanoplasmonic biosensing platform for investigating obesity-associated inflammation.

Although many advanced biosensing techniques have been proposed for cytokine profiling, there are no clinically available methods that integrate high-resolution immune cell monitoring and in situ multiplexed cytokine detection together in a biomimetic tissue microenvironment. The primary challenge arises due to the lack of suitable label-free sensing techniques and difficulty for sensor integration. In this work, we demonstrated a novel integration of a localized-surface plasmon resonance (LSPR)-based biosensor with a biomimetic microfluidic 'adipose-tissue-on-chip' platform for an in situ label-free, high-throughput and multiplexed cytokine secretion analysis of obese adipose tissue. Using our established adipose-tissue-on-chip platform, we were able to monitor the adipose tissue initiation, differentiation, and maturation and simulate the hallmark formation of crown-like structures (CLSs) during pro-inflammatory stimulation. With integrated antibody-conjugated LSPR barcode sensor arrays, our platform enables simultaneous multiplexed measurements of pro-inflammatory (IL-6 and TNF-α) and anti-inflammatory (IL-10 and IL-4) cytokines secreted by the adipocytes and macrophages. As a result, our adipose-tissue-on-chip platform is capable of identifying stage-specific cytokine secretion profiles from a complex milieu during obesity progression, highlighting its potential as a high-throughput preclinical readout for personalized obesity treatment strategies.