We report a contraction-expansion array (CEA) microchannel device that performs label-free high-throughput separation of cancer cells from whole blood at low Reynolds number (Re). The CEA microfluidic device utilizes hydrodynamic field effect for cancer cell separation, two kinds of inertial effects: (1) inertial lift force and (2) Dean flow, which results in label-free size-based separation with high throughput. To avoid cell damages potentially caused by high shear stress in conventional inertial separation techniques, the CEA microfluidic device isolates the cells with low operational Re, maintaining high-throughput separation, using nondiluted whole blood samples (hematocrit ~45%). We characterized inertial particle migration and investigated the migration of blood cells and various cancer cells (MCF-7, SK-BR-3, and HCC70) in the CEA microchannel. The separation of cancer cells from whole blood was demonstrated with a cancer cell recovery rate of 99.1%, a blood cell rejection ratio of 88.9%, and a throughput of 1.1 × 10(8) cells/min. In addition, the blood cell rejection ratio was further improved to 97.3% by a two-step filtration process with two devices connected in series.

BackgroundBiomarkers play a key role in risk assessment, assessing treatment response, and detecting recurrence and the investigation of multiple biomarkers may also prove useful in accurate prediction and prognosis of cancers. Immunohistochemistry (IHC) has been a major diagnostic tool to identify therapeutic biomarkers and to subclassify breast cancer patients. However, there is no suitable IHC platform for multiplex assay toward personalized cancer therapy. Here, we report a microfluidics-based multiplexed IHC (MMIHC) platform that significantly improves IHC performance in reduction of time and tissue consumption, quantification, consistency, sensitivity, specificity and cost-effectiveness.Methodology/Principal FindingsBy creating a simple and robust interface between the device and human breast tissue samples, we not only applied conventional thin-section tissues into on-chip without any additional modification process, but also attained perfect fluid control for various solutions, without any leakage, bubble formation, or cross-contamination. Four biomarkers, estrogen receptor (ER), human epidermal growth factor receptor 2 (HER2), progesterone receptor (PR) and Ki-67, were examined simultaneously on breast cancer cells and human breast cancer tissues. The MMIHC method improved immunoreaction, reducing time and reagent consumption. Moreover, it showed the availability of semi-quantitative analysis by comparing Western blot. Concordance study proved strong consensus between conventional whole-section analysis and MMIHC (n = 105, lowest Kendall's coefficient of concordance, 0.90). To demonstrate the suitability of MMIHC for scarce samples, it was also applied successfully to tissues from needle biopsies.Conclusions/SignificanceThe microfluidic system, for the first time, was successfully applied to human clinical tissue samples and histopathological diagnosis was realized for breast cancers. Our results showing substantial agreement indicate that several cancer-related proteins can be simultaneously investigated on a single tumor section, giving clear advantages and technical advances over standard immunohistochemical method. This novel concept will enable histopathological diagnosis using numerous specific biomarkers at a time even for small-sized specimens, thus facilitating the individualization of cancer therapy.

This work demonstrates a novel microfluidic in vitro cultivation system for embryos that improves their development using a partially constricted channel that mimics peristaltic muscle contraction. Conventional photolithography and a PDMS replica molding process were used to make straight or constricted microchannels. To investigate the effects of constriction geometry on embryonic development, different constriction widths of the channel were designed. Bovine embryos were loaded and incubated by simply placing them on a tilting machine to provide embryo movement via gravity. The fertilized embryos were cultivated on the microfluidic in vitro cultivation system until the blastocyst, hatching, or hatched blastocyst stages. To confirm the quality of blastocysts in the microfluidic channel, double staining was performed and compared with bovine embryos cultivated by the conventional droplet method. The proportion of eight-cell development among total embryos in the constricted channel (56.7+/-13.7%; mean+/-SD) was superior to that in the straight channel (23.9+/-11.0%). This suggests that the effect of constriction is vital for the early development of bovine embryos in assisted-reproduction research.

Microalgae, a group of microorganisms that grow using sunlight as the sole energy source and carbon dioxide as an only carbon source, have been considered as a feedstock of choice for the production of biofuels such as biodiesel. To explore the economic feasibility of such application, however, many technical hurdles must first be overcome; the selection and/or screening of competent species are some of the most important and yet challenging tasks. To greatly accelerate this rather slow and laborious step, we developed a droplet-based microfluidic system that uses alginate hydrogel microcapsules with a mean diameter of 26 μm, each of which is able to encapsulate a single microalgal cell. This novel device was successfully demonstrated using three microalgae species, namely, Chlorella vulgaris , Chlamydomonas sp., and Botryococcus braunii . In situ analysis of the lipid content of individual microalgal cells by nondestructive fluorescence staining using BODIPY (4,4-difluoro-1,3,5,7,-tetramethyl-4-bora-3a,4a-diaza-s-indacene) was possible. In all cases, we confirmed that the lipid content of microalgal species in alginate hydrogel microcapsules was comparable to that of free-living cells. Stochastic heterogeneity in the lipid content was verified under a highly viable physiological condition, implying that other analyses were possible after the determination of lipid content. Furthermore, the designed microwell arrays enabled us to distinguish the BODIPY fluorescence response of a single live alga within the microcapsules.

This Letter describes a quantitative phase microscopy for microfluidic devices using a simple selfreferencing interferometry. Compared with the gross dimensions of the microfluidic device, the microchannel occupies only a small area of the device. Hence, the reference field can be generated by inverting the relative position of the specimen and background. Our system is realized using an extended depth-of-field optics in the form of Michelson interferometry, which allows quantitative phase measurement for an increased depthof-field without moving objective lens or specimen. Furthermore, the system can be readily converted to a higher signal-to-noise ratio Hilbert phase microscopy thanks to the simultaneous acquisition of double interferograms. The performance of our system is verified using polymer beads, micropatterning poly(dimethylsiloxane) (PDMS), and embryo cells in the microchannels.