Nucleus of Circulating Tumor Cell Determines Its Translocation Through Biomimetic Microconstrictions and Its Physical Enrichment by Microfiltration.

The mechanism of cells passing through microconstrictions, such as capillaries and endothelial junctions, influences metastasis of circulating tumor cells (CTCs) in vivo, as well as size-based enrichment of CTCs in vitro. However, very few studies observe such translocation of microconstrictions in real time, and thus the inherent biophysical mechanism is poorly understood. In this study, a multiplexed microfluidic device is fabricated for real-time tracking of cell translocation under physiological pressure and recording deformation of the whole cell and nucleus, respectively. It is found that the deformability and size of the nucleus instead of the whole cell dominate cellular translocation through microconstrictions under a normal physiological pressure range. More specifically, cells with a large and stiff nucleus are prone to be blocked by relatively small constrictions. The same phenomenon is also observed in the size-based enrichment of CTCs from peripheral blood of metastatic cancer patients. These findings are different from a popular viewpoint that the size and deformability of a whole cell mainly determine cell translation through microconstrictions, and thus may elucidate interactions between CTCs and capillaries from a new perspective and guide the rational design of size-based microfilters for rare cell enrichment.

Evaluating a novel dimensional reduction approach for mechanical fractionation of cells using a tandem flexible micro spring array (tFMSA).

We present a novel methodology to establish experimental models for the rational design of cell fractionation based on physical properties of cells. Label-free microfluidic separation of cells based on size is a widely employed technique. However, close observation reveals that cell capture results cannot be explained by cell sizes alone. This is particularly apparent with viable cell fractionation, where cells retain their native deformability. We have developed a principal size cutoff (PSC) model based on the analysis of size distribution and size-based filtration efficiency for cell populations. The goal of this analysis is to use an unbiased approach to achieve dimensional reduction of deformability and other mechanical properties that affect cell capture. The PSC model provides a single calibrated principal size component that may be compared directly to device gap width, which is the critical dimension for cell filtration. The PSC model was evaluated experimentally using a tandem flexible micro spring array (tFMSA) device made of parylene filtration elements applied within micro-molded polydimethylsiloxane (PDMS) chambers. In the tFMSA device, a mixture of cells is sequentially passed through individual filters with decreasing gap widths to allow size-based selection. We applied this method to demonstrate viable separation of subgroups of cells with different mechanical properties from complex mixtures, including fractionation according to cancer cell type, cell cycle stage, cell viability status, and leukocyte nuclear phenotype. The PSC methodology and tFMSA device can advance a better understanding of complex factors affecting mechanical cell fractionation and provide a miniature platform for obtaining rationally designed cell fractions for biomedical applications.

[Lemierre syndrome revealed by torticollis]. / Syndrome de Lemierre révélé par un torticolis.

Classical Lemierre syndrome is a rare and severe disease with thrombosis of the internal jugular vein and metastatic infections. We report on a case of Lemierre-like syndrome secondary to mastoiditis, with a favorable outcome, in a healthy infant presenting with torticollis. Early diagnosis and treatment with antibiotics are necessary to decrease mortality.

Circulating tumor cell isolation during resection of colorectal cancer lung and liver metastases: a prospective trial with different detection techniques.

BACKGROUND: Colorectal cancer (CRC) metastasectomy improves survival, however most patient develop recurrences. Circulating tumor cells (CTCs) are an independent prognostic marker in stage IV CRC. We hypothesized that CTCs can be enriched during metastasectomy applying different isolation techniques. METHODS: 25 CRC patients undergoing liver (16 (64%)) or lung (9 (36%)) metastasectomy were prospectively enrolled (clinicaltrial.gov identifier: NCT01722903). Central venous (liver) or radial artery (lung) tumor outflow blood (7.5 ml) was collected at incision, during resection, 30 min after resection, and on postoperative day (POD) 1. CTCs were quantified with 1. EpCAM-based CellSearch® system and 2. size-based isolation with a novel filter device (FMSA). CTCs were immunohistochemically identified using CellSearch®'s criteria (cytokeratin 8/18/19+, CD45- cells containing a nucleus (DAPI+)). CTCs were also enriched with a centrifugation technique (OncoQuick®). RESULTS: CTC numbers peaked during the resection with the FMSA in contrast to CellSearch® (mean CTC number during resection: FMSA: 22.56 (SEM 7.48) (p = 0.0281), CellSearch®: 0.87 (SEM ± 0.44) (p = 0.3018)). Comparing the 2 techniques, CTC quantity was significantly higher with the FMSA device (range 0-101) than CellSearch® (range 0-9) at each of the 4 time points examined (P < 0.05). Immunofluorescence staining of cultured CTCs revealed that CTCs have a combined epithelial (CK8/18/19) and macrophage (CD45/CD14) phenotype. CONCLUSIONS: Blood sampling during CRC metastasis resection is an opportunity to increase CTC capture efficiency. CTC isolation with the FMSA yields more CTCs than the CellSearch® system. Future studies should focus on characterization of single CTCs to identify targets for molecular therapy and immune escape mechanisms of cancer cells.

Separable bilayer microfiltration device for viable label-free enrichment of circulating tumour cells.

The analysis of circulating tumour cells (CTCs) in cancer patients could provide important information for therapeutic management. Enrichment of viable CTCs could permit performance of functional analyses on CTCs to broaden understanding of metastatic disease. However, this has not been widely accomplished. Addressing this challenge, we present a separable bilayer (SB) microfilter for viable size-based CTC capture. Unlike other single-layer CTC microfilters, the precise gap between the two layers and the architecture of pore alignment result in drastic reduction in mechanical stress on CTCs, capturing them viably. Using multiple cancer cell lines spiked in healthy donor blood, the SB microfilter demonstrated high capture efficiency (78-83%), high retention of cell viability (71-74%), high tumour cell enrichment against leukocytes (1.7-2 × 10(3)), and widespread ability to establish cultures post-capture (100% of cell lines tested). In a metastatic mouse model, SB microfilters successfully enriched viable mouse CTCs from 0.4-0.6 mL whole mouse blood samples and established in vitro cultures for further genetic and functional analysis. Our preliminary studies reflect the efficacy of the SB microfilter device to efficiently and reliably enrich viable CTCs in animal model studies, constituting an exciting technology for new insights in cancer research.

Frequency resolved optical gating characterization of sub-ps pulses from single-section InAs/InP quantum dash based mode-locked lasers.

Mode-locking of single-section Fabry-Pérot InAs/InP edge emitting quantum dash based lasers at 1.56 µm under continuous wave operation is studied by second-harmonic generation frequency resolved optical gating. Self-starting pulses of a width down to 374 fs can be observed after external chirp compensation using standard single-mode fiber (SMF-28). Pulse compression using different lengths of SMF-28 and pulse shape as well as phase dependence on bias conditions are investigated. Consistency with stepped-heterodyne technique for advanced pulse characterization is shown.

Flexible micro spring array device for high-throughput enrichment of viable circulating tumor cells.

BACKGROUND: The dissemination of circulating tumor cells (CTCs) that cause metastases in distant organs accounts for the majority of cancer-related deaths. CTCs have been established as a cancer biomarker of known prognostic value. The enrichment of viable CTCs for ex vivo analysis could further improve cancer diagnosis and guide treatment selection. We designed a new flexible micro spring array (FMSA) device for the enrichment of viable CTCs independent of antigen expression. METHODS: Unlike previous microfiltration devices, flexible structures at the micro scale minimize cell damage to preserve viability, while maximizing throughput to allow rapid enrichment directly from whole blood with no need for sample preprocessing. Device performance with respect to capture efficiency, enrichment against leukocytes, viability, and proliferability was characterized. CTCs and CTC microclusters were enriched from clinical samples obtained from breast, lung, and colorectal cancer patients. RESULTS: The FMSA device enriched tumor cells with 90% capture efficiency, higher than 10(4) enrichment, and better than 80% viability from 7.5-mL whole blood samples in <10 min on a 0.5-cm(2) device. The FMSA detected at least 1 CTC in 16 out of 21 clinical samples (approximately 76%) compared to 4 out of 18 (approximately 22%) detected with the commercial CellSearch® system. There was no incidence of clogging in over 100 tested fresh whole blood samples. CONCLUSIONS: The FMSA device provides a versatile platform capable of viable enrichment and analysis of CTCs from clinically relevant volumes of whole blood.

Circulating tumor cell enrichment based on physical properties.

The metastatic dissemination and spread of malignant circulating tumor cells (CTCs) accounts for more than 90% of cancer-related deaths. CTCs detach from a primary tumor, travel through the circulatory system, and then invade and proliferate in distant organs. The detection of CTCs from blood has been established for prognostic monitoring and is predictive of patient outcome. Analysis of CTCs could enable the means for early detection and screening in cancer, as well as provide diagnostic access to tumor tissues in a minimally invasive way. The fundamental challenge with analyzing CTCs is the fact that they occur at extremely low concentrations in blood, on the order of one out of a billion cells. Various technologies have been proposed to isolate CTCs for enrichment. Here we focus on antigen-independent approaches that are not limited by specific capture antibodies. Intrinsic physical properties of CTCs, including cell size, deformability, and electrical properties, are reviewed, and technologies developed to exploit them for enrichment from blood are summarized. Physical enrichment technologies are of particular interest as they have the potential to increase yield and enable the analysis of rare CTC phenotypes that may not be otherwise obtained.

23 and 39 GHz low phase noise monosection InAs/InP (113)B quantum dots mode-locked lasers.

Here we report for the first time a passive mode-locking of single section Fabry-Perot (FP) lasers based on InAs quantum dots(QDs) grown on (113)B InP substrate. Devices under study are a 1 and 2 mm long laser diodes emitting around 1.58 µm. Self-starting pulses with repetition rates around 23 and 39 GHz and pulse widths down to 1.5 ps are observed after propagation through a suitable length of single-mode fiber for intracavity dispersion compensation. A RF spectral width as low as 20 kHz has been obtained leading to a low timing jitter RMS.

Viable circulating tumor cell enrichment by flexible micro spring array.

We demonstrated a high throughput versatile platform capable of isolating circulating tumor cells (CTCs) from clinically relevant volumes of blood while preserving their viability and ability to proliferate. The enrichment is based on the fact that CTCs are larger compared with normal blood cells. The incorporated system allows size-based separation of CTCs at the micro-scale, while taking advantage of a high throughput and rapid processing speed. Testing results of model systems using cell lines show that this device can enrich CTCs from 7.5 mL of whole blood samples with 90% capture efficiency, higher than 10(4) enrichment, and better than 80% viability in approximately ten minutes without any incidence of clogging.

Single step measurement of optical transmitters Henry factor using sinusoidal optical phase modulations.

Measurement of the Henry factor over large optical bandwidth is carried out in a single step without any filtering, using a technique based on the sinusoidal phase modulation method. This fast technique was successfully applied to a directly modulated Fabry Perot laser to obtain simultaneously the linewidth enhancement factor (LEF) of 14 longitudinal modes. It is also well suited for electro-absorption modulators (EAM) for which the α-factor is determined over 15 nm optical bandwidth. A very good agreement is found with the well established fiber transfer function method.

Excitable phase slips in an injection-locked single-mode quantum-dot laser.

An experimental study of the dynamics of a single-mode quantum-dot semiconductor laser undergoing optical injection is described for the first time, to our knowledge. In particular, the first observation of excitable pulses near the locking boundaries for both positive and negative detuning is reported, indicating locking via a saddle-node bifurcation for both signs of the detuning. The phase evolution of the slave electric-field during pulsing was measured and confirmed that the pulses result from 2pi phase slips. The interpulse-time statistics were analyzed, and a Kramers-like distribution was obtained.

Experimental investigation of the timing jitter in self-pulsating quantum-dash lasers operating at 1.55 microm.

We report for the first time on the systematic measurement of timing jitter of 40-GHz self-pulsating Fabry-Perot laser based on InAs/InP quantum dashes emitting at 1.55 microm. Two different methods, one based on optical cross-correlation and one on electrical spectrum sideband integration are used and show a good agreement, yielding a jitter of 0.86 ps in the 1 MHz---20 MHz frequency range with a potential of 280 fs for optimized driving conditions. Amplitude noise and high-frequency timing jitter contributions are also discussed.

Short pulse generation using a passively mode locked single InGaAsP/InP quantum well laser.

We report on subpicosecond pulse generation using passively mode locked lasers (MLL) based on a low optical confinement single InGaAsP/InP quantum well active layer grown in one epitaxial step. Systematic investigation of the performances of two-section MLLs emitting at 1.54 microm evidenced pulse width of 860 fs at 21.31 GHz repetition rate, peak power of approximately 500 mW and a time-bandwith product of 0.57. A 30 kHz linewidth of the photodetected radio-frequency electrical spectrum is further demonstrated at 21 GHz which is, to our knowledge, the lowest value ever reported for a quantum well device.

Coherence collapse and low-frequency fluctuations in quantum-dash based lasers emitting at 1.57 mum.

Optical feedback tolerance is experimentally investigated on a 600-mum-long quantum-dash based Fabry-Pérot laser emitting at 1.57mum. While quantum-dashes are structurally intermediate to quantum-wells and quantum-dots, the observed behaviour is distinctly like that of a quantum-well based laser but with greater stability. Coherence collapse and low-frequency fluctuation regimes are observed and are reported here. The onset of the coherence collapse regime is experimentally determined and is found to vary from -29 dB to -21 dB external feedback level when increasing the current from twice to nine times the threshold current.

Phase-amplitude characterization of a high-repetition-rate quantum dash passively mode-locked laser.

We apply a novel phase-amplitude characterization method to a one-section quantum dash-based passively mode-locked laser at a 42.2 GHz repetition rate. The method relies on the measurement of the spectral phase of the longitudinal modes by the successive analysis of the correlation signal of a group of three adjacent modes. It provides both the temporal shape of the intensity and the phase of the emitted signal. A pulse of 1.5 ps of width is measured, and a pedestal is exhibited. Extinction ratio limitation is explained by investigating the origin of this pedestal. The accuracy of the method is estimated by comparing the measured autocorrelation signal and the calculated one from the phase analysis.