Microfluidic Immiscible Phase Filtration System for the Isolation of Small Numbers of Cells from Whole Blood

Abstract

Isolation of circulating tumor cells (CTCs) has generated clinical and academic interest due to the important role that CTCs play in cancer metastasis and diagnosis. Here, we present a PDMS and glass prototype of a microfluidic device for the immunomagnetic, immiscible phase filtration based capture, and isolation of MCF-7 breast cancer cells, from various sample matrices including PBS-based buffer, blood plasma, and unprocessed whole blood. Following optimization of surface energy of an oil-water interface, microfluidic geometry, and bead-binding kinematics, our microfluidic device achieved 95 AE 4% recovery of target cells from PBS-based buffer with 95% purity, 90 AE 3% recovery of target cells from blood plasma and recovery of~70 AE 5% from unprocessed whole blood with purity >99% with 1 ml blood samples with 1,000 spiked target cells. From quantitative studies to assess the nonspecific carryover of contaminants from whole blood, we found that our system accomplishes a >175 fold depletion in platelets, >900 fold depletion in erythrocytes, and >1,700 fold depletion in leukocytes with respect to unprocessed whole blood, enabling us to avoid sample pre-processing. In addition, we found that~95% of the isolated target cells were viable, making them suitable for subsequent molecular and cellular studies. We quantify and propose mechanisms for the carryover of platelet, erythrocyte, and leukocyte contamination in purified samples, rather than relying on sample pre-processing. These results validate the continued study of our platform for extraction of CTCs from patient samples and other rare cell isolation applications.

Advances in Biosensors and Diagnostic Technologies Using Nanostructures and Nanomaterials

“…[247,248] Additionally, antibody-functionalized superparamagnetic beads have been used to isolate rare cells from whole blood samples on microfluidic platforms. [249,250] Poudineh et al developed a unique microfluidic chip architecture that separates circulating tumor cells (CTCs) based on the level of interaction of antibody-functionalized magenetic beads within these cells. This device enabled phentoypic profiling of the CTCs in samples as dilute as 10 CTCs per mL of blood.…”

Preparation of Tissues and Heterogeneous Cellular Samples for Single-Cell Analysis

“…Specifically bound cells are then removed from solution using a magnet, followed by washing of the beads and liberation of the bound cells. Common cellular-recognition elements include the Epithelial Cell Adhesion Molecule (EPCAM) for separation of CTCs from whole blood, such as is used in the commercially available Veridex Cellsearch system and other applications [39]. Other approaches can use separation via specifically engineered indirect interactions, such as biotin/streptavidin coupling.…”