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Markovitz-Bishitz Y.,Bar - Ilan University | Tauber Y.,Bar - Ilan University | Afrimzon E.,Bar - Ilan University | Zurgil N.,Bar - Ilan University | And 5 more authors.

Multicellular spheroid models have been recognized as superior to monolayer cell cultures in antitumor drug screening, but their commercial adaptation in the pharmaceutical industry has been delayed, primarily due to technological limitations. The current study presents a new spheroid culture platform that addresses these technical restrictions. The new culturing device is based on a multiwell plate equipped with a glass bottom patterned with an array of UV adhesive microchambers. Each microchamber is designed to accommodate a single spheroid. The system facilitates the simultaneous creation and culturing of a large number of spheroids, as well as screening their response to antitumor drugs. The volume of the spheroids is easily controlled by seeding density. The location of each spheroid is preserved in the same microchamber throughout its growth, treatment with soluble agents, and imaging. The growth ratio parameter, a non-intrusive size analysis of the same spheroid before and after exposure to drugs, was found to be a sensitive indicator for the reaction of MCF7 breast cancer spheroids to cytotoxic drugs. This feature helps reveal the heterogeneity within the spheroid population during the formation process and their drug response, and provides an opportunity to detect specific, highly active or drug-resistant spheroid sub-groups. The advantages of this spheroid-based system make it an efficient drug-screening tool that may be valuable to related fields of research and clinical applications. © 2010 Elsevier Ltd. Source

Kohler S.,University of Leipzig | Weilbeer C.,University of Leipzig | Howitz S.,GeSiM mbH | Becker H.,Microfluidic ChipShop GmbH | And 2 more authors.
Lab on a Chip - Miniaturisation for Chemistry and Biology

In this work, a microfluidic free-flow electrophoresis device with a novel approach for preventing gas bubbles from entering the separation area is presented. This is achieved by integrating partitioning bars to reduce the channel depth between electrode channels and separation chamber in order to obtain electrical contact and simultaneously prevent bubbles from entering the separation area. The three-layer sandwich chip features a reusable carrier plate with integrated ports for fluidic connection combined with a softlithographically cast microfluidic PDMS layer and a sealing glass slide. This design allows for a straightforward and rapid chip prototyping process. The performance of the device is demonstrated by free-flow zone electrophoretic separations of fluorescent dye mixtures as well as by the separation of labeled amines and amino acids with separation voltages up to 297 V. © 2011 The Royal Society of Chemistry. Source

Afrimzon E.,Bar - Ilan University | Zurgil N.,Bar - Ilan University | Shafran Y.,Bar - Ilan University | Ehrhart F.,Fraunhofer Institute for Biomedical Engineering | And 11 more authors.
BMC Cell Biology

Background: The cryopreservation and thawing processes are known to induce many deleterious effects in cells and might be detrimental to several cell types. There is an inherent variability in cellular responses among cell types and within individual cells of a given population with regard to their ability to endure the freezing and thawing process. The aim of this study was to evaluate the fate of cryopreserved cells within an optical cryo apparatus, the individual-cell-based cryo-chip (i3C), by monitoring several basic cellular functional activities at the resolution of individual cells.Results: In the present study, U937 cells underwent the freezing and thawing cycle in the i3C device. Then a panel of vital tests was performed, including the number of dead cells (PI staining), apoptotic rate (Annexin V staining), mitochondrial membrane potential (TMRM staining), cytoplasm membrane integrity and intracellular metabolism (FDA staining), as well as post-thawing cell proliferation assays. Cells that underwent the freezing - thawing cycle in i3C devices exhibited the same functional activity as control cells. Moreover, the combination of the multi-parametric analysis at a single cell resolution and the optical and biological features of the device enable an accurate determination of the functional status of individual cells and subsequent retrieval and utilization of the most valuable cells.Conclusions: The means and methodologies described here enable the freezing and thawing of spatially identifiable cells, as well as the efficient detection of viable, specific, highly biologically active cells for future applications. © 2010 Afrimzon et al; licensee BioMed Central Ltd. Source

Deutsch M.,Bar - Ilan University | Afrimzon E.,Bar - Ilan University | Namer Y.,Bar - Ilan University | Shafran Y.,Bar - Ilan University | And 10 more authors.
BMC Cell Biology

Background: Cryopreservation is the only widely applicable method of storing vital cells for nearly unlimited periods of time. Successful cryopreservation is essential for reproductive medicine, stem cell research, cord blood storage and related biomedical areas. The methods currently used to retrieve a specific cell or a group of individual cells with specific biological properties after cryopreservation are quite complicated and inefficient.Results: The present study suggests a new approach in cryopreservation, utilizing the Individual Cell-based Cryo-Chip (i3C). The i3C is made of materials having appropriate durability for cryopreservation conditions. The core of this approach is an array of picowells, each picowell designed to maintain an individual cell during the severe conditions of the freezing - thawing cycle and accompanying treatments. More than 97% of cells were found to retain their position in the picowells throughout the entire freezing - thawing cycle and medium exchange. Thus the comparison between pre-freezing and post-thawing data can be achieved at an individual cell resolution. The intactness of cells undergoing slow freezing and thawing, while residing in the i3C, was found to be similar to that obtained with micro-vials. However, in a fast freezing protocol, the i3C was found to be far superior.Conclusions: The results of the present study offer new opportunities for cryopreservation. Using the present methodology, the cryopreservation of individual identifiable cells, and their observation and retrieval, at an individual cell resolution become possible for the first time. This approach facilitates the correlation between cell characteristics before and after the freezing - thawing cycle. Thus, it is expected to significantly enhance current cryopreservation procedures for successful regenerative and reproductive medicine. © 2010 Deutsch et al; licensee BioMed Central Ltd. Source

Kohler S.,University of Leipzig | Becker H.,Microfluidic ChipShop GmbH | Beushausen V.,Laser Laboratorium Gottingen E.V. | Huttner W.,Laser Laboratorium Gottingen E.V. | And 4 more authors.
Proceedings of SPIE - The International Society for Optical Engineering

In this work we present the first approach towards low-cost free-flow electrophoresis (FFE) devices utilizing injection molding as a microfabrication process which has the potential to manufacture FFE chips at a cost which make their use commercially viable. This is achieved by realizing a new straightforward micro free-flow electrophoresis (μFFE) design ensuring both, bubble free electrophoretic separation and effective electrical connection by implementing miniaturized partitioning bars. This creates a defined open gap of 20 μm in height and 500 μm in width between separation zone and electrode channels. The thermoplastic μFFE chips are ready to use, there is no need for a subsequent labor-intensive implementation of membranes or salt bridges to separate the electrode channels from the separation zone. © 2011 Copyright Society of Photo-Optical Instrumentation Engineers (SPIE). Source

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