Demonstrates efficient large-scale isolation of milk exosomes, suggesting a breakthrough in biological particle separation technology.
Free‐flow electrophoresis is a core technology for biological particle separation, but traditional devices suffer from low automation, poor flow stability, and limited throughput, hindering industrial‐scale applications. Herein, a novel fully automatic reciprocating free‐flow isoelectric focusing (RFFIEF) system, a batch‐type of automated closed‐loop free‐flow isoelectric focusing (FFIEF), was developed, integrating a 24‐channel separation chamber, closed gas–liquid circulation drive, and intelligent control system to achieve unattended standardized operation. Standard protein verification showed excellent flow stability and pH gradient reproducibility (pooled standard deviation [SD] = 0.19 pH units excluding ampholyte‐induced outliers). The system was applied to large‐scale separation of milk exosomes, which were effectively focused at pH 5.0–7.5. Exosome subpopulations were fractionated by surface charge: CD81‐positive subpopulations concentrated at pH 5.8–6.5, whereas CD63‐positive subpopulations were highly enriched at pH 6.9. This system breaks through the automation bottleneck of conventional devices, providing an efficient platform for large‐scale biological particle separation with broad industrial prospects.
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Wang et al. (2026) studied this question.
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