Microfluidics guided by deep learning for cancer immunotherapy screening.

Immunocyte infiltration and cytotoxicity play critical roles in both inflammation and immunotherapy. However, current cancer immunotherapy screening methods overlook the capacity of the T cells to penetrate the tumor stroma, thereby significantly limiting the development of effective treatments for solid tumors. Here, we present an automated high-throughput microfluidic platform for simultaneous tracking of the dynamics of T cell infiltration and cytotoxicity within the 3D tumor cultures with a tunable stromal makeup. By recourse to a clinical tumor-infiltrating lymphocyte (TIL) score analyzer, which is based on a clinical data-driven deep learning method, our platform can evaluate the efficacy of each treatment based on the scoring of T cell infiltration patterns. By screening a drug library using this technology, we identified an epigenetic drug (lysine-specific histone demethylase 1 inhibitor, LSD1i) that effectively promoted T cell tumor infiltration and enhanced treatment efficacy in combination with an immune checkpoint inhibitor (anti-PD1) in vivo. We demonstrated an automated system and strategy for screening immunocyte-solid tumor interactions, enabling the discovery of immuno- and combination therapies.

Trapping cell spheroids and organoids using digital acoustofluidics.

The precise positioning and arrangement of cell spheroids and organoids are critical to reconstructing complex tissue architecture for tissue engineering and regenerative medicine. Here, we present a digital acoustofluidic method to manipulate cell spheroids and organoids with unprecedented dexterity. By introducing localized vibrations via a C-shaped integrated digital transducer (IDT), we can generate a trapping node to immobilize cell spheroids with a diameters ranging from 20µm to 300µm. Moreover, we digitally trapped multiple cell spheroids atop the C-shaped IDTs within a closed or open microfluidic chamber. By programming the trapping nodes within a 3 × 3 C-shaped IDT array, we can precisely position cell spheroids into designed patterns. We also demonstrated that our digital acoustofluidic device can accurately control the interaction of spheroid cells and organoids. Along with a simple fabrication procedure and setup, our digital acoustofluidic method can provide precisely manipulate and position various cell spheroids or organoids in a contactless, label-free, and highly biocompatible manner. We believe this technology can be widely used for tissue engineering, regenerative medicine, and fundamental cell biology research.

Profiling Cell-Matrix Adhesion Using Digitalized Acoustic Streaming.

Profiling the kinetics of cell-matrix adhesion is of great importance to understand many physiological and pathological processes such as morphogenesis, tissue homeostasis, wound healing, and tumorigenesis. Here, we developed a novel digital acoustofluidic device for parallel profiling cell-matrix adhesion at single-cell level. By introduction of localized and uniform acoustic streaming into an open chamber microfluidic device, the adherent cells within the open chamber can be detached by the streaming-induced Stokes drag force. By digital regulation of pulsed acoustic power from a low level to high levels, the hundreds of adherent cells can be ruptured from the fibronectin-coated substrate accordingly, and their adhesive forces (from several pN to several nN) and kinetics can be determined by the applied power and cell incubation time. As a proof-of-concept application for studying cancer metastasis, we applied this technique to measure the adhesion strength and kinetics of human breast cancer cells to extracellular matrix such as fibronectin and compared their metastatic potentials by measuring the rupture force of cancer cells representing malignant (MCF-7 cells and MDA-MB-231 cells) and nonmalignant (MCF-10A cells) states. Our acoustofluidic device is simple, easy to operate, and capable of measuring, in parallel, hundreds of individual cells' adhesion forces with a resolution at the pN level. Thus, we expect this device could be widely used for both fundamental cell biology research as well as development of cancer diagnostics and tissue engineering technologies.

A Digital Acoustofluidic Pump Powered by Localized Fluid-Substrate Interactions.

The precise transportation of small-volume liquids in microfluidic and nanofluidic systems remains a challenge for many applications, such as clinical fluidical analysis. Here, we present a reliable digital pump that utilizes acoustic streaming induced by localized fluid-substrate interactions. By locally generating streaming via a C-shaped interdigital transducer (IDT) within a triangle-edged microchannel, our acoustofluidic pump can generate a stable unidirectional flow (∼nanoliter per second flow rate) with a precise digital regulation (∼second response time), and it is capable of handling aqueous solutions (e.g., PBS buffer) as well as high viscosity liquids (e.g., human blood) with a nanoliter-scale volume. Along with our acoustofluidic pump's low cost, programmability, and capacity to control small-volumes at high precision, it could be widely used for point-of-care diagnostics, precise drug delivery, and fundamental biomedical research.

High-throughput acoustofluidic fabrication of tumor spheroids.

Three-dimensional (3D) culture of multicellular spheroids, offering a desirable biomimetic microenvironment, is appropriate for recapitulating tissue cellular adhesive complexity and revealing a more realistic drug response. However, current 3D culture methods are suffering from low-throughput, poor controllability, intensive-labor, and variation in spheroid size, thus not ready for many high-throughput screening applications including drug discovery and toxicity testing. Herein, we developed a high-throughput multicellular spheroid fabrication method using acoustofluidics. By acoustically-assembling cancer cells with low-cost and disposable devices, our method can produce more than 12 000 multicellular aggregates within several minutes and allow us to transfer these aggregates into ultra-low attachment dishes for long-term culture. This method can generate more than 6000 tumor spheroids per operation, and reduce tumor spheroid formation time to one day. Our platform has advantages in forming spheroids with high throughput, short time, and long-term effectiveness, and is easy-to-operation. This acoustofluidic spheroid assembly method provides a simple and efficient way to produce large numbers of uniform-sized spheroids for biomedical applications in translational medicine, pharmaceutical industry and basic life science research.