Khan, M. Z. U. | Search bioRxiv

bioRxiv2025

CellTrap: A Microfluidic Platform Enabling Cell-Cell Interactions at Variable Effector to Target Ratios

Immune-cancer cell interactions play a central role in understanding antitumor responses and evaluating immunotherapies. However, long-term, single-cell-level analysis of these interactions remains challenging. To address this, we developed a microfluidic trapping device with 1,024 traps, each equipped with a filter to retain cells, sustain medium flow, minimize cross-talk, and allow precise control of effector-to-target (E:T) ratios. The platform enables continuous monitoring of immune-cancer interactions for up to 14 hours. Device characterization was performed using 10 {micro}m fluorescent beads seeded via hydrostatic flow, with trap occupancy validated by Poisson statistics. Initial experiments using PBMCs against GFP-expressing U87 (U87GFP) glioblastoma cells demonstrated an immune-mediated reduction in GFP intensity, which was interpreted cautiously as a cytotoxic response. To improve reproducibility, we subsequently employed IL-2-stimulated Natural Killer cells (NK92IL2) as standardized effectors and evaluated their interactions with U87GFP glioblastoma cells, K562 chronic myelogenous leukemia cells, and LS174T adenocarcinoma cells. Time-lapse imaging revealed transient intracellular calcium fluxes, consistent with early activation of NK92IL2 cells, followed by a cytotoxic response. Increasing E:T ratios consistently enhanced immune activity, highlighting the utility of this device for dissecting immune-cancer interactions and guiding the development of immunotherapy.

bioengineering↗

bioRxiv2025

3D Hydrodynamic Flow Lithography

Continuous- and stop-flow lithography has been widely used for the fabrication of ingenious multi-dimensional microstructures, including fibers and microparticles. The flow profile of multiple co-flowing streams during the process, as one of the dimensions, dictates the cross-sectional morphology of the microstructures. Here, we introduce a three-dimensional hydrodynamic flow lithography (3D HFL) approach that enables rapid and programmable generation of desired flow profiles and their conversion into solid microstructures. By strategically combining multiple flow sculpting strategies, including variable inlet configurations, intra-channel pillar configurations, and outlet configurations, we achieved precise and versatile control over flow sculpting, as validated by computational fluid dynamics simulations and experimental production of microparticles and fibers, offering unparalleled design flexibility. Importantly, the flow sculpting device is fabricated using low-cost 3D-printed molds to cast PDMS channels with precisely aligned inlets and complex geometries that are difficult or impossible to achieve using conventional soft lithography. Our 3D HFL approach effectively overcomes the limitations of existing microfabrication techniques in device complexity, structural diversity, multi-material integration, and production throughput. Furthermore, we demonstrated the capability of this platform to produce anisotropic multi-material microparticles and fibers tailored for specific applications, such as amphiphilic particles for uniform microdroplet capture, biocompatible patterned hydrogels for controlled cell adhesion, and dual-layer fibers for temperature sensing. The simplicity of device fabrication, combined with the broad design flexibility, establishes this platform as a scalable, high-throughput, and versatile solution for engineering anisotropic microparticles and fibers with tailored functionalities, providing powerful tools for a wide range of downstream fields.

bioengineering↗