Transfection microarray of human mesenchymal stem cells and on-chip siRNA gene knockdown

Yoshikawa, Tomohiro; Uchimura, Eiichiro; Kishi, Michiko; Funeriu, Daniel P.; Miyake, Masato; Miyake, Jun

doi:10.1016/j.jconrel.2004.01.024

Cited by 130 publications

(102 citation statements)

References 11 publications

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“…Yoshikawa et al achieved localized GFP knock-down in human mesenchymal stem cells on arrayed spots of transfection reagent, reporter plasmids, siRNAs and fibronectin 9 . Expanding the compatibility of cell microarrays to nonadherent and semiadherent cell lines, Kato et al designed a biocompatible anchor for membrane surface to fabricate slides amenable to the attachment of nonadherent human erythroleukemic K562 cells 11 .…”

Section: Rnai Cell Microarraysmentioning

confidence: 99%

“…Given the apparatus necessary for screens in 96-or 384-well plates, high-throughput loss-of-function screening would be simplified by the use of cell microarrays for large RNAi experiments [7][8][9][10][11][12][13][14][15] . RNAi cell microarray slides should hold at least 5,000-6,000 spots of RNAi reagents, enabling whole-genome screens on a small number of slides 8,15 .…”

Section: Cell Microarrays As a Vehicle For High-throughput Rnai Screensmentioning

confidence: 99%

“…Efforts by a number of groups to develop cell microarrays using RNAi have introduced the possibility of using cell microarrays for largescale RNAi studies [7][8][9][10][11][12][13][14][15] . Although this technique is still in its infancy, cell microarrays can considerably reduce the amount of effort necessary for rapid cell-based RNAi screens.…”

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confidence: 99%

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Cell microarrays and RNA interference chip away at gene function

2005

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S25 P E R S P E C T I V EThe recent development of cell microarrays offers the potential to accelerate high-throughput functional genetic studies. The widespread use of RNA interference (RNAi) has prompted several groups to fabricate RNAi cell microarrays that make possible discrete, in-parallel transfection with thousands of RNAi reagents on a microarray slide. Though still a budding technology, RNAi cell microarrays promise to increase the efficiency, economy and ease of genome-wide RNAi screens in metazoan cells.

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Section: Rnai Cell Microarraysmentioning

confidence: 99%

Section: Cell Microarrays As a Vehicle For High-throughput Rnai Screensmentioning

confidence: 99%

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Cell microarrays and RNA interference chip away at gene function

2005

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“…Also, DNA complexes have been mixed with fibronectin and spotted onto glass slides for arrayed transfection of human mesenchymal stem cells [51]. While transfected cell arrays hold great potential [46][47][48][49][50][51], further development of a substratemediated delivery system that efficiently transfects a wide variety of primary cells and cell lines, while allowing for spatially-controlled delivery within the different domains is required [46,52,53]. Using SAMs on gold to form transfected cell arrays can allow precise control over surface chemistries, interactions between the substrate and DNA complexes, transfection efficiencies, and pattern sizes to create a well-characterized and efficient array delivery system.…”

Section: Patterned Transfectionmentioning

confidence: 99%

Substrate-mediated delivery from self-assembled monolayers: Effect of surface ionization, hydrophilicity, and patterning

2005

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Gene transfer has many potential applications in basic and applied sciences. In vitro, DNA delivery can be enhanced by increasing the concentration of DNA in the cellular microenvironment through immobilization of DNA to a substrate that supports cell adhesion. Substrate-mediated delivery describes the immobilization of DNA, complexed with cationic lipids or polymers, to a biomaterial or substrate. As surface properties are critical to the efficiency of the surface delivery approach, selfassembled monolayers (SAMs) of alkanethiols on gold were used to correlate surface chemistry of the substrate to binding, release, and transfection of non-specifically immobilized complexes. Surface hydrophobicity and ionization were found to mediate both DNA complex immobilization and transfection, but had no effect on complex release. Additionally, SAMs were used in conjunction with soft lithographic techniques to imprint substrates with specific patterns, resulting in patterned DNA complex deposition and transfection, with transfection efficiencies in the patterns nearing 40%. Controlling the interactions between complexes and substrates, with the potential for patterned delivery, can be used to locally enhance or regulate gene transfer, with applications to tissue engineering scaffolds and transfected cell arrays.

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“…Gene delivery from the culture surface, termed substrate-mediated delivery (Segura and Shea, 2002;Segura et al, 2003), reverse transfection (Ziauddin and Sabatini, 2001), or solid-phase delivery (Bielinska et al, 2000), is being developed to enhance gene transfer and to create transfected cell arrays (Yoshikawa et al, 2004;Ziauddin and Sabatini, 2001). Several strategies have been developed to associate DNA complexes with the substrate including specific binding of complexes to substrate through the biotin-avidin interaction (Segura and Shea, 2002;Segura et al, 2003), gelatin entrapment (Ziauddin and Sabatini, 2001), or nonspecific adsorption (Yoshikawa et al, 2004). These systems have shown the capacity to transfect cells cultured on the substrate, with deposition by adsorption transfecting the greatest number of cell types.…”

Section: Introductionmentioning

confidence: 99%

Gene delivery through cell culture substrate adsorbed DNA complexes

Bengali

Pannier

Segura

et al. 2005

Biotech & Bioengineering

130

187

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Efficient gene delivery is a fundamental goal of biotechnology and has numerous applications in both basic and applied science. Substrate-mediated delivery and reverse transfection enhance gene transfer by increasing the concentration of DNA in the cellular microenvironment through immobilizing a plasmid to a cell culture substrate prior to cell seeding. In this report, we examine gene delivery of plasmids that were complexed with cationic polymers (polyplexes) or lipids (lipoplexes) and subsequently immobilized to cell culture or biomaterial substrates by adsorption. Polyplexes and lipoplexes were adsorbed to either tissue culture polystyrene or serumadsorbed tissue culture polystyrene. The quantity of DNA immobilized increased with time of exposure, and the deposition rate and final amount deposited depended upon the properties of the substrate and complex. For polyplexes, serum modification enhanced reporter gene expression up to 1500-fold relative to unmodified substrates and yielded equivalent or greater expression compared to bolus delivery. For lipoplexes, serum modification significantly increased the number of transfected cells relative to unmodified substrates yet provided similar levels of expression. Immobilized complexes transfect primary cells with improved cellular viability relative to bolus delivery. Finally, this substrate-mediated delivery approach was extended to a widely used biomaterial, poly(lactide-coglycolide). Immobilization of DNA complexes to tissue culture polystyrene substrates can be a useful tool for enhancing gene delivery for in vitro studies. Additionally, adapting this system to biomaterials may facilitate application to fields such as tissue engineering.

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Transfection microarray of human mesenchymal stem cells and on-chip siRNA gene knockdown

Cited by 130 publications

References 11 publications

Cell microarrays and RNA interference chip away at gene function

Cell microarrays and RNA interference chip away at gene function

Substrate-mediated delivery from self-assembled monolayers: Effect of surface ionization, hydrophilicity, and patterning

Gene delivery through cell culture substrate adsorbed DNA complexes

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