Functional Stem Cell Sorting via Integrative Droplet Synchronization

Abstract

Stem cell regenerative medicine strategy requires selecting functional cells to trigger repair processes. Stem cell secretion measurement is important to evaluate cellular activities for functional cell sorting. At present, to determine single cell secretions, mixing chemical sensors and cells together in a chamber is a standard procedure. However, toxic chemical sensors, such as albumin assay kits, are used during this process, causing low viability (64%) and low functionality (30%). It is especially important for stem cell profiling, as the toxicity of chemical sensors such as albumin permanently changes stem cell phenotypes, leading to unwanted analysis outcomes. Moreover, because of the sensor toxicity, the challenge of culturing sorted cells remain. In this study, an integrative synchronized droplet screen system was developed to separate a large droplet with cell encapsulation into two daughter droplets: one droplet containing cell secretions and the other droplet containing a single cell. These two daughter droplets moved along the channels at the same speed in synchronization. By injecting toxic chemical sensors into one daughter droplet, the single-cell secretions were determined without affecting the cells in the corresponding droplet. Based on the daughter droplet synchronization, the cells without mixing toxicity sensors were sorted for cell culturing. For example, to identify hepatocytes, the albumin secretion of undifferentiated HepaRG stem cells was measured in daughter droplets by injecting a toxic albumin assay kit for functional stem cell sorting. With synchronized sorting, functional hepatocytes were collected without exposure to toxic chemical sensors, showing high viability (78%) and active functionality (89%).

Microfluidic Platforms for Real‐Time In Situ Monitoring of Biomarkers for Cellular Processes

“…However, they have certain limitations. Currently, microfluidic platforms for cell analysis rely heavily on fluorescence technology, [155][156]159,161,178] and there is a need to develop cell screening and analysis techniques based on cell characteristics such as size, hydrophilicity, and polarity.…”

“…Sun et al developed a microfluidic platform based on fluorescence for functional stem cell sorting that exhibited a high sorting efficiency of 90% (Figure 10a). [159] In another study, Wang et al designed a microfluidic platform combined with machine learning for tumor cell classification. [120] As shown in Figure 10b, the authors optimized the cell separation technology and investigated the effects of material modification.…”

Recent Advances in Microfluidic Technology for Bioanalysis and Diagnostics

“…Chemical sensors can be harmful to cells and may induce phenotype changes, especially for stem cells. To overcome this bottleneck, a droplet-based platform was recently presented, which splits single-cell droplets for secretion analysis (Figure A) . Single stem cells were first encapsulated into droplets and produced albumin.…”

“…To overcome this bottleneck, a droplet-based platform was recently presented, which splits single-cell droplets for secretion analysis (Figure 6A). 157 Single stem cells were first encapsulated into droplets and produced albumin. After splitting the droplets, a cell-toxic albumin sensor was added to the daughter droplet without the cell and analyzed by fluorescence spectroscopy.…”

Smart Droplet Microfluidic System for Single-Cell Selective Lysis and Real-Time Sorting Based on Microinjection and Image Recognition

“…Passive sorting methods mainly work based on size sorting, which is related to deterministic lateral displacement (DLD). , Although passive sorting methods are easy to operate and process, they are less precise and adaptable than active methods. Active single-cell droplet sorting can be categorized as electrical, magnetic, acoustic, , and hydrodynamic sorting methods, depending on the detailed external fields applied. Compared with passive sorting, active methods rely on physical factors to achieve higher accuracy, flexibility, and throughput.…”