MagSculptor: A Microfluidic Platform for High-Resolution Magnetic Fractionation of Low-Expression Cell Subtypes.

Heterogeneous expression of a single surface protein within one cell population can drive major functional differences, yet low-expression subtypes remain difficult to isolate. Conventional tube-based immunomagnetic separation collapses all labelled cells into one positive fraction and thus cannot resolve small differences in marker abundance. Here, we present MagSculptor, a microfluidic platform for high-resolution magnetic fractionation of low-expression EpCAM-defined subtypes within one immunomagnetically labelled population at a time. Arrays of soft-magnetic strips create localized high-gradient zones that map modest differences in bead loading onto distinct capture positions, yielding High (H), Medium (M), Low (L), and Negative (N) fractions. Finite element method simulations of coupled magnetic and hydrodynamic fields quantify the field gradients and define an operating window. Experimentally, epithelial cancer cell lines processed sequentially under identical settings show reproducible subtype partitioning. In a low-EpCAM model (MDA-MB-231), conventional flow cytometry, under standard EpCAM staining conditions, did not yield a robust EpCAM-positive gate, whereas MagSculptor still revealed graded subpopulations. Western blotting confirms a monotonic decrease in EpCAM abundance from H to N, and doxorubicin assays show distinct in vitro drug sensitivities, while viability remains above 95%. MagSculptor thus helps extend immunomagnetic separation from binary enrichment to multi-level isolation of low-expression subtypes and provides a convenient front-end for downstream functional and molecular analyses.