Circulating tumor cells (CTCs) have been linked to cancer progression but are difficult to isolate, as they are very rare and heterogeneous, covering a range of sizes and expressing different molecular receptors. Filtration has emerged as a simple and powerful method to enrich CTCs but only captures cells above a certain size regardless of molecular characteristics. Here, we introduce antibody-functionalized microfilters to isolate CTCs based on both size and surface receptor expression. We present a 3D printed filtration cartridge with microfabricated polymer filters with 8, 10, 12, 15, or 20 μm-diameter pores. Pristine filters were used to optimize sample dilution, rinsing protocol, flow rate, and pore size, leading to >80% for the recovery of spiked cancer cells with very low white blood cell contamination (<1000). Then, filters were functionalized with antibodies against either epithelial cell adhesion molecule (EpCAM) or epidermal growth factor receptor (EGFR) and the cartridges were used to enrich breast (MDA-MB-231, MCF-7) and renal (786-O, A-498) cancer cells expressing various levels of EpCAM and EGFR. Cancer cells were spiked into human blood, and when using filters with antibodies specific to a molecular receptor expressed on a cell, efficiency was increased to >96%. These results suggest that filtration can be optimized to target specific CTC characteristics such as size and receptor expression and that a diverse range of CTCs may be captured using particular combinations of pore size, filtration parameters, and antibody functionalization.
A vacuum assisted UV micro-molding (VAUM) process is proposed for the fabrication of freestanding and defect-free polymer membranes based on a UV-curable methacrylate polymer (MD 700). VAUM is a highly flexible and powerful method for fabricating low cost, robust, large-area membranes over 9 × 9 cm with pore sizes from 8 to 20 μm in diameter, 20 to 100 μm in thickness, high aspect ratio (the thickness of the polymer over the diameter of the hole is up to 15 : 1), high porosity, and a wide variety of geometrical characteristics. The fabricated freestanding membranes are flexible while mechanically robust enough for post manipulation and handling, which allows them to be cut and integrated as a plastic cartridge onto thermoplastic 3D microfluidic devices with single or double filtration stages. Very high particle capture efficiencies (≈98%) have been demonstrated in the microfluidic devices integrated with polymer membranes, even when the size of the beads is very close to the size of the pores of the microfilter. About 85% of the capture efficiency has been achieved in cancer cell trapping experiments, in which a breast cancer cell line (MDA-MB-231) spiked with phosphate-buffered saline buffer when the pore size of the filter is 8 μm and the device is operated at a flow rate of 0.1 mL min.
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