The CryoCapsule: simplifying correlative light to electron microscopy.

Correlating complementary multiple scale images of the same object is a straightforward means to decipher biological processes. Light microscopy and electron microscopy are the most commonly used imaging techniques, yet despite their complementarity, the experimental procedures available to correlate them are technically complex. We designed and manufactured a new device adapted to many biological specimens, the CryoCapsule, that simplifies the multiple sample preparation steps, which at present separate live cell fluorescence imaging from contextual high-resolution electron microscopy, thus opening new strategies for full correlative light to electron microscopy. We tested the biological application of this highly optimized tool on three different specimens: the in vitro Xenopus laevis mitotic spindle, melanoma cells over-expressing YFP-langerin sequestered in organized membranous subcellular organelles and a pigmented melanocytic cell in which the endosomal system was labeled with internalized fluorescent transferrin.

Formation of UHMWPE Nanofibers during Solid-State Deformation.

A network of nanofibers is formed in situ through solid-state deformation of disentangled ultra-high molecular weight polyethylene (dis-UHMWPE) during compounding with a polyolefin elastomer below the melting temperature of dis-UHMWPE crystals. Dis-UHMWPE was prepared in the form of powder particles larger than 50 µm by polymerization at low temperatures, which favored the crystallization and prevention of macromolecules from entangling. Shearing the blend for different durations and at different temperatures affects the extent to which the grains of dis-UHMWPE powder deform into nanofibers. Disentangled powder particles could deform into a network of nanofibers with diameters between 110 and 340 nm. The nanocomposite can be further sheared for a longer time to decrease the diameter of dis-UHMWPE nanofibers below 40 nm, being still composed of ≈70 wt.% of crystalline and ≈30 wt.% of amorphous components. Subsequently, these thinner fibers begin to melt and fragment because they are thinner and also because the amorphous defects locally decrease the nanofibers' melting temperature, which results in their fragmentation and partial loss of nanofibers. These phenomena limit the thickness of dis-UHMWPE nanofibers, and this explains why prolonged or more intense shearing does not lead to thinner nanofibers of dis-UHMWPE when compounded in a polymeric matrix.

A mesofluidic multiplex immunosensor for detection of circulating cytokeratin-positive cells in the blood of breast cancer patients.

We have recently reported the analytical performance of an immunosensor comprising one mm-scale parallel plate laminar flow chamber and applied to capture MCF7 breast cancer cells (Ehrhart et al., Biosens. Bioelectr. 24, 467, 2008). Herein we present a new multiplex immunosensor embodying four parallel plate laminar flow chambers that fit onto a standard, functionalized, microscopy glass slide. The four surfaces are coated with long alkyl chain spacers of 21-aminohenicosyl trichlorosilane (AHTS) and then grafted with a monoclonal anti-human epithelial cell adhesion molecule (EpCAM) antibody specific of target cells to immobilize. We first demonstrate a significantly (P < 0.01) improved capacity of each of the four flow chambers of the multiplex immunosensor to capture MCF7 cells compared to the previous single chamber device. Second, in addition to an increase of cell immobilization, the multiplex device offers a versatile tool easily grafted with various purified antibodies onto the four surfaces. Third, we obtained high cell capture rate and efficiency of various numbers of MCF7 cells spiked in buffer containing an equal number of background leukocytes. And fourth, we demonstrate isolation efficiency of circulating tumor cells (CTCs) from peripheral blood drawn from a small cohort of patients with localized or metastatic breast cancer. This new multiplex immunosensor could be tested for its potential to capture different subpopulations of CTCs.

Fiber Orientation and Concentration in an Injection-Molded Ethylene-Propylene Copolymer Reinforced by Hemp.

This paper characterizes and analyzes the microstructures of injection-molded polypropylene parts reinforced with 20 wt% of hemp fibers in order to understand the process induced variations in thermomechanical properties. In-thickness fiber orientation and fiber content were determined by X-ray tomography along the flow. The fiber content along the flow path was also determined by direct fiber content measurements after matrix dissolution, showing an increase of 2%/100 mm for a 2.2 mm-thick plate due to fiber migration during the filling stage. A typical shell/core structure for fiber orientation in injection molding was observed, but with a very clear transition between the layer solidified under high shear rates and the core in which the fiber content was reduced by more than 50%. The orientation of hemp fibers is lower than the one of glass fibers, especially in thickness direction. However, the overall fiber orientation in the injection direction induces significant anisotropic thermomechanical properties, which cannot be explained by simple micromechanical models that consider isotropic mechanical properties for hemp fibers. These phenomena must be taken into account in process simulation codes for injection molding to better predict thermomechanical properties as well as part shrinkage and warpage to design molds.

Impact of Nanoconfinement on Polylactide Crystallization and Gas Barrier Properties.

The barrier properties of poly(l-lactide) (PLLA) were investigated in multinanolayer systems, probing the effect of confinement, the compatibility between the confining and the confined polymer, crystal orientation, and amorphous phase properties. The multilayer coextrusion process was used to confine PLLA between two amorphous polymers (polystyrene, PS; and polycarbonate, PC), which have different chemical affinities with PLLA. Confined PLLA layers of approximately 20 nm thickness were obtained. The multinanolayer materials were annealed at different temperatures to obtain PLLA crystallites with distinct polymorphs. PLLA annealed in PC/PLLA films at 120 °C afforded a crystallinity degree up to 65%, and PLLA annealed in PC/PLLA or PS/PLLA films at 85 °C had a crystallinity degree of 45%. WAXS measurements evidenced that the PLLA lamellas between PS layers had a mixed in-plane and on-edge orientation. PLLA lamellas between PC layers were uniquely oriented in-plane. DMA results evidenced a shift of the PC glass transition toward lower temperature, suggesting the possible presence of an interphase. The development of the rigid amorphous fraction (RAF) in the amorphous phase during annealing was impacted by the confiner polymer. The RAF content of semicrystalline PLLA was about 15% in PC/PLLA, whereas it was neglectable in PS/PLLA. The oxygen barrier properties appeared to be governed by RAF content, and no impact of the PLLA polymorph or the crystalline orientation was observed. This study shows that the confinement of PLLA on itself does not impact barrier properties but that the proper choice of the confiner polymer can lead to decrease the phase coupling which creates the RAF. It is the prevention of RAF that decreases permeability.

Prediction and Sensitivity Analysis of Bubble Dissolution Time in 3D Selective Laser Sintering Using Ensemble Decision Trees.

The presence of defects like gas bubble in fabricated parts is inherent in the selective laser sintering process and the prediction of bubble shrinkage dynamics is crucial. In this paper, two artificial intelligence (AI) models based on Decision Trees algorithm were constructed in order to predict bubble dissolution time, namely the Ensemble Bagged Trees (EDT Bagged) and Ensemble Boosted Trees (EDT Boosted). A metadata including 68644 data were generated with the help of our previously developed numerical tool. The AI models used the initial bubble size, external domain size, diffusion coefficient, surface tension, viscosity, initial concentration, and chamber pressure as input parameters, whereas bubble dissolution time was considered as output variable. Evaluation of the models' performance was achieved by criteria such as Mean Absolute Error (MAE), Root Mean Squared Error (RMSE) and coefficient of determination (R2). The results showed that EDT Bagged outperformed EDT Boosted. Sensitivity analysis was then conducted thanks to the Monte Carlo approach and it was found that three most important inputs for the problem were the diffusion coefficient, initial concentration, and bubble initial size. This study might help in quick prediction of bubble dissolution time to improve the production quality from industry.

Preparation and characterization of poly(ethylene terephthalate) films coated by chitosan and vermiculite nanoclay.

Chitosan (CS) layers are coated on a poly(ethylene terephthalate) (PET) film in order to decrease the oxygen permeability through the polymeric films for food packaging applications. Oxygen transmission rate (OTR) of the 130 µm PET films can be decreased from 11 to only 0.31 cm3/m².day with a coated layer of 2 µm of CS. Additional decrease is obtained with the addition of vermiculite (VMT) to CS matrix in high proportion (40 to 50 w/w%). The OTR of the coated PET films decreased to very low values, below the detection limit of commercial instrumentation (≤0.008 cm3/m2 day). This high-barrier behavior is believed to be due to the brick wall nanostructure, which produces an extremely tortuous path for oxygen molecules.