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Circulating rare cells (CRCs) in the peripheral blood are considered crucial cells in pathological phenomena. Circulating tumor cells (CTCs) detach from solid tumors via blood transport and play a crucial role in cancer metastasis, which is the primary focus of CRCs research. Therefore, a simple and cost-effective device for capturing and culturing CTCs is required for pathological analysis. This study proposes a new biosensing device that includes a pair of surface-modified pico-porous nanostructured chips with a large cross-section and contact area. Compared to traditional narrow microfluidics, this study features a lower shear stress force, resulting in less damage to the captured cells (less than 5%). In addition, unlike devices with non-nanostructured silicon chips, this microcavity biosensing device can provide approximately ten times the cell capture rate and maintain the captured cells as living cells. Another important point to note is that captured live cells can be cultured for up to 18 days or longer. Computer-aided cell image recognition was utilized to minimize human misjudgment and to reduce the analysis time to less than 30 minutes. The novelty of this research is the improved capture rate of CTCs and the prolonged survival time of cells in culture achieved through the utilization of the newly proposed device.

Abstract Background Noninvasive prenatal testing (NIPT) based on cell-free DNA in maternal circulation has been accepted worldwide by the clinical community since 2011 but limitations, such as maternal malignancy and fetoplacental mosaicism, preclude its full replacement of invasive prenatal diagnosis. We present a novel silicon-based nanostructured microfluidics platform named as “Cell Reveal™” to demonstrate the feasibility of capturing circulating fetal nucleated red blood cells (fnRBC) and extravillous cytotrophoblasts (EVT) for cell-based noninvasive prenatal diagnosis (cbNIPD). Methods The “Cell Reveal™” system is a silicon-based, nanostructured microfluidics using immunoaffinity to capture the trophoblasts and the nucleated RBC (nRBC) with specific antibodies. The automated computer analysis software was used to identify the targeted cells through additional immunostaining of the corresponding antigens. The identified cells were retrieved for whole genome amplification for subsequent investigations by micromanipulation in one microchip, and left in situ for subsequent fluorescence in situ hybridization (FISH) in another microchip. When validation, bloods from pregnant women (n = 24) at gestational age 11–13+6 weeks were enrolled. When verification, bloods from pregnant women (n = 5) receiving chorionic villus sampling or amniocentesis at gestation age 11+4–21 weeks with an aneuploid or euploid fetus were enrolled, followed by genetic analyses using FISH, short tandem repeat (STR) analyses, array comparative genomic hybridization, and next generation sequencing, in which the laboratory is blind to the fetal genetic complement. Results The numbers of captured targeted cells were 1–44 nRBC/2 ml and 1–32 EVT/2 ml in the validation group. The genetic investigations performed in the verification group confirmed the captured cells to be fetal origin. In every 8 ml of the maternal blood being blindly tested, both fnRBC and EVT were always captured. The numbers of captured fetal cells were 14–22 fnRBC/4 ml and 1–44 EVT/4 ml of maternal blood. Conclusions This report is one of the first few to verify the capture of fnRBC in addition to EVT. The scalability of our automated system made us one step closer toward the goal of in vitro diagnostics.

Epithelial ovarian cancer (EOC) is a leading cause of cancer mortality among women but unfortunately is usually not diagnosed until advanced stage. Early detection of EOC is of paramount importance to improve outcomes. Liquid biopsy of circulating tumor cells (CTCs) is emerging as one of the promising biomarkers for early detection of solid tumors. However, discrepancies in terms of oncogenomics (i.e., different genetic defects detected) between the germline, primary tumor, and liquid biopsy are a serious concern and may adversely affect downstream cancer management. Here, we illustrate the potential and pitfalls of CTCs by presenting two patients of Stage I EOC. We successfully isolated and recovered CTCs by a silicon-based nanostructured microfluidics system, the automated Cell RevealTM. We examined the genomics of CTCs as well as the primary tumor and germline control (peripheral blood mononuclear cells) by whole exome sequencing. Different signatures were then investigated by comparisons of identified mutation loci distinguishing those that may only arise in the primary tumor or CTCs. A novel model is proposed to test if the highly variable allele frequencies, between primary tumor and CTCs results, are due to allele dropout in plural CTCs or tumor heterogeneity. This proof-of-principle study provides a strategy to elucidate the possible cause of genomic discrepancy between the germline, primary tumor, and CTCs, which is helpful for further large-scale use of such technology to be integrated into clinical management protocols.

Circulating fetal cells (CFCs) in maternal blood are rare but have a strong potential to be the target for noninvasive prenatal diagnosis (NIPD). “Cell RevealTM system„ is a silicon-based microfluidic platform capable to capture rare cell populations in human circulation. The platform is recently optimized to enhance the capture efficiency and system automation. In this study, spiking tests of SK-BR-3 breast cancer cells were used for the evaluation of capture efficiency. Then, peripheral bloods from 14 pregnant women whose fetuses have evidenced non-maternal genomic markers (e.g., de novo pathogenic copy number changes) were tested for the capture of circulating fetal nucleated red blood cells (fnRBCs). Captured cells were subjected to fluorescent in situ hybridization (FISH) on chip or recovered by an automated cell picker for molecular genetic analyses. The capture rate for the spiking tests is estimated as 88.1%. For the prenatal study, 2⁻71 fnRBCs were successfully captured from 2 mL of maternal blood in all pregnant women. The captured fnRBCs were verified to be from fetal origin. Our results demonstrated that the Cell RevealTM system has a high capture efficiency and can be used for fnRBC capture that is feasible for the genetic diagnosis of fetuses without invasive procedures.