The ability to isolate and analyze rare circulating tumor cells (CTCs) has the potential to further our understanding of cancer metastasis and enhance the care of cancer patients. In this protocol, we describe the procedure for isolating rare CTCs from blood samples by using tumor antigen–independent microfluidic CTC-iChip technology. The CTC-iChip uses deterministic lateral displacement, inertial focusing and magnetophoresis to sort up to 107 cells/s. By using two-stage magnetophoresis and depletion antibodies against leukocytes, we achieve 3.8-log depletion of white blood cells and a 97% yield of rare cells with a sample processing rate of 8 ml of whole blood/h. The CTC-iChip is compatible with standard cytopathological and RNA-based characterization methods. This protocol describes device production, assembly, blood sample preparation, system setup and the CTC isolation process. Sorting 8 ml of blood sample requires 2 h including setup time, and chip production requires 2–5 d.

When Segré and Silberberg in 1961 witnessed particles in a laminar pipe flow congregating at an annulus in the pipe, scientists were perplexed and spent decades learning why such behavior occurred, finally understanding that it was caused by previously unknown forces on particles in an inertial flow. The advent of microfluidics opened a new realm of possibilities for inertial focusing in the processing of biological fluids and cellular suspensions and created a field that is now rapidly expanding. Over the past five years, inertial focusing has enabled high-throughput, simple, and precise manipulation of bodily fluids for a myriad of applications in point-of-care and clinical diagnostics. This review describes the theoretical developments that have made the field of inertial focusing what it is today and presents the key applications that will make inertial focusing a mainstream technology in the future.

This report details a comprehensive study of inertial focusing dynamics and particle behavior in low aspect ratio (h/w ∼ 1/1 to 1/8) spiral microchannels. A continuum of particle streak behavior is shown with longitudinal, cross-sectional, and velocity resolution, yielding a large analyzed parameter space. The dataset is then summarized and compared to prior results from both straight microchannels and other low aspect ratio spiral microchannel designs. Breakdown of focusing into a primary and secondary fluorescent streak is observed in the lowest aspect ratio channels at high average downstream velocities. Streak movement away from the theoretically predicted near inner wall equilibrium position towards the center of the channel at high average downstream velocities is also detailed as a precursor to breakdown. State diagrams detail the overall performance of each device including values of the required channel lengths and the range of velocities over which quality focusing can be achieved.