Nanosecond Pulsed Laser Activated Massively Parallel Single-cell Intracellular Delivery Using Ti Micro-Dish
27 Sep 2020-pp 320-324
TL;DR: The proposed device platform is an array of photothermal Micro-dish patterned with fibronectin protein to obtain specific Single-cell attachment and can deliver 10,000 individual cells with 97% efficiency and 95% cell viability simultaneously.
Abstract: Single-cell intracellular delivery is a booming field of interest with the various challenges faced due to the impermeability of cargo to the cell membrane barrier. The idea is to locally disrupt cell membrane for allowing cargo passage into the cytosol such as to create its therapeutic effect. Till to date, most of the intracellular delivery is performed in bulk approaches, which provides an average ensemble data, losing low-frequency rare information. Besides, reliable results require huge data from multiple cells. Hence, to ensure huge data on individual cells it becomes necessary to carry out delivery experiments on a large number of single-cells simultaneously. This ensures the same experimental conditions conserving heterogeneity in the cell population. The collection of data from individual cells provides information about rare low-frequency alleles in the cell population. Massively parallel delivery ensures such uniform experimental conditions with uniform cargo delivery to a large number of individual cells. The proposed device platform is an array of photothermal Micro-dish patterned with fibronectin protein to obtain specific Single-cell attachment. The device can deliver 10,000 individual cells with 97% efficiency and 95% cell viability simultaneously.
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TL;DR: Time-resolved imaging was used to examine the use of pulsed laser microbeam irradiation to produce cell lysis and indicated that cells subject to transient shear stresses in excess of a critical value were lysed while cells exposed to lowerShear stresses remained adherent and viable.
197 citations
"Nanosecond Pulsed Laser Activated M..." refers background in this paper
...An increase in the pulse energy increases the spatial extent of the impact of photothermal bubbles [7]....
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TL;DR: In this review, this review of the recent advances in single-cell technologies and their applications insingle-cell manipulation, diagnosis, and therapeutics development are described.
Abstract: The investigation of human disease mechanisms is difficult due to the heterogeneity in gene expression and the physiological state of cells in a given population. In comparison to bulk cell measurements, single-cell measurement technologies can provide a better understanding of the interactions among molecules, organelles, cells, and the microenvironment, which can aid in the development of therapeutics and diagnostic tools. In recent years, single-cell technologies have become increasingly robust and accessible, although limitations exist. In this review, we describe the recent advances in single-cell technologies and their applications in single-cell manipulation, diagnosis, and therapeutics development.
63 citations
"Nanosecond Pulsed Laser Activated M..." refers background in this paper
...Two major challenges faced during SCT studies are one the sensitivity of the measuring instrument should be high enough to detect the low quantities of the analyte present in control as well as sample and second is obtaining huge data of statistical significance from multiple experiments to give reliable conclusions [2]...
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TL;DR: Single-cell analysis at the threshold is described as a novel and scalable approach to single-cell cell analysis that aims to provide real-time information about the architecture of the cell and its response to treatment.
Abstract: 1111 Xi Chen, Michael J.T. Stubbington and Sarah A. Teichmann are at the Wellcome Trust Sanger Institute, Cambridge, UK; J. Christopher Love is at the Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA; Nicholas E. Navin is in the Department of Genetics and Department of Bioinformatics and Computational Biology, The University of Texas, MD Anderson Cancer Center, Houston, Texas, USA; Lior Pachter is in the Department of Mathematics, University of California, Berkeley, California, USA; Valentine Svensson is at the European Bioinformatics Institute, European Molecular Biology Laboratory, Cambridge, UK; and Jonathan V. Sweedler is in the Department of Chemistry, University of Illinois, Urbana, Illinois, USA. e-mail: lpachter@math.berkeley.edu, sweedler@scs.illinois.edu, clove@mit.edu, nnavin@mdanderson.org or sarah.teichmann@sanger.ac.uk Single-cell analysis at the threshold
61 citations
"Nanosecond Pulsed Laser Activated M..." refers background in this paper
...Further, cell lineage can be traced to multiple generations with the ease of live cell imaging [4]....
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TL;DR: In infrared (IR) pulse laser-activated highly efficient parallel intracellular delivery by using an array of titanium microdish (TMD) device, which is compact, easy-to-use, and potentially applicable for cellular therapy and diagnostic purposes.
Abstract: We report infrared (IR) pulse laser-activated highly efficient parallel intracellular delivery by using an array of titanium microdish (TMD) device. Upon IR laser pulse irradiation, a two-dimensional array of TMD device generated photothermal cavitation bubbles to disrupt the cell membrane surface and create transient membrane pores to deliver biomolecules into cells by a simple diffusion process. We successfully delivered the dyes and different sizes of dextran in different cell types with variations of laser pulses. Our platform has the ability to transfect more than a million cells in a parallel fashion within a minute. The best results were achieved for SiHa cells with a delivery efficiency of 96% and a cell viability of around 98% for propidium iodide dye using 600 pulses, whereas a delivery efficiency of 98% and a cell viability of 100% were obtained for dextran 3000 MW delivery using 700 pulses. For dextran 10,000 MW, the delivery efficiency was 92% and the cell viability was 98%, respectively. The device is compact, easy-to-use, and potentially applicable for cellular therapy and diagnostic purposes.
22 citations
"Nanosecond Pulsed Laser Activated M..." refers background in this paper
...This shows the non-toxic nature of the photoporation platform [6]....
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01 Jan 2020
TL;DR: This chapter mainly focuses on different physical drug-delivery techniques such as electroporation, optoporation, mechanopsoration, magnetoporation and hybrid techniques along with their working mechanisms, advantages, disadvantages, and limitations.
Abstract: Delivery of exogenous materials or cargo such as drugs, proteins, peptides, and nucleic acids into cells is a vital segment in molecular and cellular biology for potential cellular therapy and drug-discovery applications contributing toward personalization of medicine. Over the years, drug-delivery techniques have been developed in order to gain more control over the drug dosage, targeted delivery, and to minimize side effects. The major drug-delivery techniques can be classified as viral, chemical, and physical methods. Viral vectors are prominently used for gene therapy; however, they are cell-specific and have an immune response with high toxicity. Chemical methods are often limited by the low efficiency of plasmid delivery into different cell types due to plasmid degradation and toxicity. Considering these limitations, different physical methods such as photoporation, gene gun, hydrodynamic injection, electroporation, and mechanoporation, etc., are being widely developed for highly efficient cargo delivery with low toxicity. These methods are able to create transient hydrophilic membrane pores to deliver cargos into cells using different physical energies. Currently, ex vivo cargo delivery is widely studied while few in vivo applications have been developed. Concerning several obstacles to cargo delivery into cells, this chapter mainly focuses on different physical drug-delivery techniques such as electroporation, optoporation, mechanoporation, magnetoporation, and hybrid techniques along with their working mechanisms, advantages, disadvantages, and limitations. An insight into the future prospects and real-time applications of these techniques is also discussed.
18 citations