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      Nanofabrication of Isoporous Membranes for Cell Fractionation

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          Abstract

          Cell fractionations and other biological separations frequently require several steps. They could be much more effectively done by filtration, if isoporous membranes would be available with high pore density, and sharp pore size distribution in the micro- and nanoscale. We propose a combination of two scalable methods, photolithography and dry reactive ion etching, to fabricate a series of polyester membranes with isopores of size 0.7 to 50 μm and high pore density with a demonstrated total area of 38.5 cm 2. The membranes have pore sizes in the micro- and submicro-range, and pore density 10-fold higher than track-etched analogues, which are the only commercially available isoporous polymeric films. Permeances of 220,000 L m −2 h −1bar −1 were measured with pore size 787 nm. The method does not require organic solvents and can be applied to many homopolymeric materials. The pore reduction from 2 to 0.7 μm was obtained by adding a step of chemical vapor deposition. The isoporous system was successfully demonstrated for the organelle fractionation of Arabidopsis homogenates and could be potentially extended to other biological fractionations.

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          Membrane microfilter device for selective capture, electrolysis and genomic analysis of human circulating tumor cells.

          This paper presents development of a parylene membrane microfilter device for single stage capture and electrolysis of circulating tumor cells (CTCs) in human blood, and the potential of this device to allow genomic analysis. The presence and number of CTCs in blood has recently been demonstrated to provide significant prognostic information for patients with metastatic breast cancer. While finding as few as five CTCs in about 7.5mL of blood (i.e., 10(10) blood cells in) is clinically significant, detection of CTCs is currently difficult and time consuming. CTC enrichment is performed by either gradient centrifugation of CTC based on their buoyant density or magnetic separation of epithelial CTC, both of which are laborious procedures with variable efficiency, and CTC identification is typically done by trained pathologists through visual observation of stained cytokeratin-positive epithelial CTC. These processes may take hours, if not days. Work presented here provides a micro-electro-mechanical system (MEMS)-based option to make this process simpler, faster, better and cheaper. We exploited the size difference between CTCs and human blood cells to achieve the CTC capture on filter with approximately 90% recovery within 10 min, which is superior to current approaches. Following capture, we facilitated polymerase chain reaction (PCR)-based genomic analysis by performing on-membrane electrolysis with embedded electrodes reaching each of the individual 16,000 filtering pores. The biggest advantage for this on-membrane in situ cell lysis is the high efficiency since cells are immobilized, allowing their direct contact with electrodes. As a proof-of-principle, we show beta actin gene PCR, the same technology can be easily extended to real time PCR for CTC-specific transcript to allow molecular identification of CTC and their further characterization.
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            Thinking the future of membranes: Perspectives for advanced and new membrane materials and manufacturing processes

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              Size-selective microcavity array for rapid and efficient detection of circulating tumor cells.

              Circulating tumor cells (CTCs) are tumor cells circulating in the peripheral blood of patients with metastatic cancer. Detection of CTCs has clinical significance in cancer therapy because it would enable earlier diagnosis of metastasis. In this research, a microfluidic device equipped with a size-selective microcavity array for highly efficient and rapid detection of tumor cells from whole blood was developed. The microcavity array can specifically separate tumor cells from whole blood on the basis of differences in the size and deformability between tumor and hematologic cells. Furthermore, the cells recovered on the microcavity array were continuously processed for image-based immunophenotypic analysis using a fluorescence microscope. Our device successfully detected approximately 97% of lung carcinoma NCI-H358 cells in 1 mL whole blood spiked with 10-100 NCI-H358 cells. In addition, breast, gastric, and colon tumor cells lines that include EpCAM-negative tumor cells, which cannot be isolated by conventional immunomagnetic separation, were successfully recovered on the microcavity array with high efficiency (more than 80%). On an average, approximately 98% of recovered cells were viable. Our microfluidic device has high potential as a tool for the rapid detection of CTCs and can be used to study CTCs in detail.
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                Author and article information

                Contributors
                suzana.nunes@kaust.edu.sa
                Journal
                Sci Rep
                Sci Rep
                Scientific Reports
                Nature Publishing Group UK (London )
                2045-2322
                9 April 2020
                9 April 2020
                2020
                : 10
                : 6138
                Affiliations
                [1 ]ISNI 0000 0001 1926 5090, GRID grid.45672.32, King Abdullah University of Science and Technology (KAUST), Biological and Environmental Science and Engineering (BESE) Division, Advanced Membranes and Porous Materials Center, ; 23955-6900 Thuwal, Saudi Arabia
                [2 ]ISNI 0000 0001 1926 5090, GRID grid.45672.32, King Abdullah University of Science and Technology (KAUST), Nanofabrication Core Laboratory, ; 23955-6900 Thuwal, Saudi Arabia
                [3 ]ISNI 0000 0001 1926 5090, GRID grid.45672.32, King Abdullah University of Science and Technology (KAUST), Biological and Environmental Science and Engineering (BESE) Division, Center for Desert Agriculture, ; 23955-6900 Thuwal, Saudi Arabia
                Author information
                http://orcid.org/0000-0003-2056-3735
                http://orcid.org/0000-0003-3119-9633
                http://orcid.org/0000-0002-3669-138X
                Article
                62937
                10.1038/s41598-020-62937-5
                7145805
                32273573
                6adf978d-b9fc-4595-8bc6-0db9c88aaee1
                © The Author(s) 2020

                Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

                History
                : 9 December 2019
                : 15 March 2020
                Categories
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                © The Author(s) 2020

                Uncategorized
                plant sciences,nanopores
                Uncategorized
                plant sciences, nanopores

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