A microfluidics-based method for isolation and visualization of cells based on receptor-ligand interactions

Receptor-ligand interactions are typically studied using biochemical assays like isothermal titration calorimetry or surface plasmon resonance. These methods provide insights at the molecular level but lack the ability to visualize interactions at the cellular level. Existing techniques also require large cell populations, making single-cell analysis difficult. Prior research has shown that gene expression changes can indicate receptor-ligand interactions, but direct visualization remains a gap. No single-cell method currently exists for directly observing these interactions. This gap motivated the development of new tools to study receptor-ligand binding at the individual cell level. Researchers have long sought ways to isolate and observe cells based on specific surface markers. The absence of a reliable single-cell visualization method has limited progress in this area. This study addresses that limitation by introducing a novel microfluidic approach.

The aim of this study is to develop a method for visualizing and isolating cells based on receptor-ligand interactions at the single-cell level. Current techniques either lack visualization capabilities or require large cell numbers. This work seeks to bridge that gap by introducing a microfluidic chip-based system. The researchers focus on isolating and observing cells that bind to immobilized ligands. They aim to demonstrate the effectiveness of this approach using T cells and FGL2. The study also tests the versatility of the method by applying it to another receptor-ligand pair. The goal is to provide a reliable and efficient protocol for single-cell analysis. This approach could enable new insights into cell signaling and function. The method is designed to be accessible for further downstream processing.

The researchers designed a microfluidic chip-based system to isolate and visualize cells based on receptor-ligand interactions. First, a protein is immobilized on a glass slide using a low-flow-rate pump. Cells are then passed through the chip to identify those expressing receptors that bind to the immobilized ligand. The study uses biotin-conjugated FGL2 immobilized on an avidin-coated slide chip. A mixture of GFP-labeled wild-type T cells and RFP-labeled FcγRIIB-knockout T cells is introduced into the system. Automated scanning and counting are used to assess binding activity. The method is further validated by isolating T cells based on their affinity to vimentin. The system allows for single-cell level analysis and isolation in about four hours.

The study found a large number of GFP+ T cells with binding activity to the immobilized FGL2. In contrast, significantly fewer RFP+ FcγRIIB-knockout T cells bound to the ligand. Automated scanning confirmed the differential binding between the two cell types. The method successfully isolated cells expressing specific receptors. The researchers further demonstrated the versatility of the system by isolating T cells expressing a membrane-anchored IL-12 variant. The binding was specific to the receptor's affinity for vimentin. The protocol enables efficient single-cell analysis within four hours. These findings suggest the method is effective for studying receptor-ligand interactions at the individual cell level.

The authors conclude that their microfluidic chip-based method enables direct visualization of receptor-ligand interactions at the single-cell level. The study demonstrates the system's ability to distinguish between cells expressing and lacking specific receptors. The method successfully isolates cells based on binding activity. The researchers confirm the versatility of the approach by applying it to a different receptor-ligand pair. The protocol allows for efficient downstream processing of isolated cells. The system's performance suggests it is suitable for further studies in receptor-ligand interactions. The authors propose that this method could be used to study a wide range of cell surface interactions. The findings support the use of this technique for single-cell analysis in immunology.