Metastasis is the main cause of tumor-related death, and the dispersal of tumor cells through the circulatory system is a critical step in the metastatic process. Early detection and analysis of circulating tumor cells (CTCs) is therefore important for early diagnosis, prognosis, and effective treatment of cancer, enabling favorable clinical outcomes in cancer patients. Accurate and reliable methods for isolating and detecting CTCs are necessary to obtain this clinical information. Over the past two decades, microfluidic technologies have demonstrated great potential for isolating and detecting CTCs from blood. The present paper reviews current advanced microfluidic technologies for isolating CTCs based on various biological and physical principles, and discusses their fundamental advantages and drawbacks for subsequent cellular and molecular assays. Owing to significant genetic heterogeneity among CTCs, microfluidic technologies for isolating individual CTCs have recently been developed. We discuss these single-cell isolation methods, as well as approaches to overcoming the limitations of current microfluidic CTC isolation technologies. Finally, we provide an overview of future innovative microfluidic platforms.

We developed an epithelial cell adhesion molecule (EpCAM)-based positive method and CD45/CD66b-based negative method for isolating circulating tumor cells (CTCs) by lateral magnetophoresis. The CTC recovery rate, white blood cell depletion rate, and purity of CTCs isolated using the positive and negative methods were analyzed using blood samples spiked with cancer cells with different expression levels of EpCAM. The aim was to assess the strengths and weaknesses of the positive and negative isolation methods for CTC-based diagnostics, prognostics, and therapeutics for cancer. The EpCAM-based positive method yielded CTCs of high purity, while the CD45/CD66b-based negative method yielded a large number of CTCs. In conclusion, the positive method shows promise for detecting somatic oncogenic mutations and the negative method shows promise for discovery of cellular and transcriptomic biomarkers of cancer.

This paper introduces a single-cell isolation technology for circulating tumor cells (CTCs) using a microfluidic device (the "SIM-Chip"). The SIM-Chip comprises a lateral magnetophoretic microseparator and a microdispenser as a two-step cascade platform. First, CTCs were enriched from whole blood by the lateral magnetophoretic microseparator based on immunomagnetic nanobeads. Next, the enriched CTCs were electrically identified by single-cell impedance cytometer and isolated as single cells using the microshooter. Using 200 μL of whole blood spiked with 50 MCF7 breast cancer cells, the analysis demonstrated that the single-cell isolation efficiency of the SIM-Chip was 82.4%, and the purity of the isolated MCF7 cells with respect to WBCs was 92.45%. The data also showed that the WBC depletion rate of the SIM-Chip was 2.5 × 10(5) (5.4-log). The recovery rates were around 99.78% for spiked MCF7 cells ranging in number from 10 to 90. The isolated single MCF7 cells were intact and could be used for subsequent downstream genetic assays, such as RT-PCR. Single-cell culture evaluation of the proliferation of MCF7 cells isolated by the SIM-Chip showed that 84.1% of cells at least doubled in 5 days. Consequently, the SIM-Chip could be used for single-cell isolation of rare target cells from whole blood with high purity and recovery without cell damage.

We describe an assembly-disposable microfluidic device based on a silicone-coated release polymer thin film. It consists of a disposable polymeric superstrate and a reusable functional substrate and they are assembled simply using vacuum pressure. The disposable polymeric superstrate is manufactured by bonding a silicone-coated release polymer thin film and a microstructured polydimethylsiloxane (PDMS) replica, containing only a simple structured microchannel. The reusable functional substrate generates an intricate energy field that can penetrate the micrometer-thick polymer film into the microchannel and control microfluids. This is the first report to introduce a silicone-coated release polyethylene terephthalate (PET) thin film as a bonding layer on a microstructured PDMS replica. The bonding strength was ∼600 kPa, which is the strongest among bonding methods of PDMS and PET polymer. Additionally, accelerated tests for bond stability and leakage demonstrated that the silicone-coated release PET film can form a very robust bond with PDMS. To demonstrate the usefulness of the proposed assembly-disposable microfluidic device, a lateral magnetophoretic microseparator was developed in an assembly-disposable microfluidic device format and was evaluated for isolating circulating tumor cells (CTCs) from patients with breast cancer.

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