Efficient propagation of single cells accutase‐dissociated human embryonic stem cells

Bajpai, Ruchi; Lesperance, Jacqueline; Kim, Min; Terskikh, Alexey V.

doi:10.1002/mrd.20809

Cited by 145 publications

(111 citation statements)

References 20 publications

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“…Second, we used Accutase (a mixture of proteolytic and collagenolytic enzymes) to dissociate the cells. It has been reported that using Accutase instead of trypsin can significantly improve cell viability (Bajpai et al 2008). Third, we treated cells with a ROCK (Rho-associated kinase) inhibitor before and during transfection.…”

Section: Resultsmentioning

confidence: 99%

High-efficiency siRNA-based gene knockdown in human embryonic stem cells

Ma¹,

Jin²,

Dong³

et al. 2010

RNA

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Loss-of-function studies in human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) via nonviral approaches have been largely unsuccessful. Here we report a simple and cost-effective method for high-efficiency delivery of plasmids and siRNAs into hESCs and iPSCs. Using this method for siRNA delivery, we achieve >90% reduction in the expression of the stem cell factors Oct4 and Lin28, and observe cell morphological and staining pattern changes, characteristics of hESC differentiation, as a result of Oct4 knockdown.

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Section: Resultsmentioning

confidence: 99%

High-efficiency siRNA-based gene knockdown in human embryonic stem cells

Ma¹,

Jin²,

Dong³

et al. 2010

RNA

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“…Other investigators have reported that recombinant vitronectin 12 , a hydrogel 13 , and a LN-511 fragment 14 can serve as alternatives to Matrigel or feeder cells for culturing of hES cells. However, none of the cell culture substrata allows clonal survival of hES cells without the use of apoptosis inhibitors [15][16][17] or chemically undefined substances 18,19 , and the cells are often passaged in small clumps that can contain a significant proportion of differentiated cells. Since the initial number of cells is low during derivation of new ES cell lines, absence of cell culture conditions that enable clonal survival also complicates the derivation procedure especially from single blastomeres.…”

mentioning

confidence: 99%

Clonal culturing of human embryonic stem cells on laminin-521/E-cadherin matrix in defined and xeno-free environment

et al. 2014

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Lack of robust methods for establishment and expansion of pluripotent human embryonic stem (hES) cells still hampers development of cell therapy. Laminins (LN) are a family of highly cell-type specific basement membrane proteins important for cell adhesion, differentiation, migration and phenotype stability. Here we produce and isolate a human recombinant LN-521 isoform and develop a cell culture matrix containing LN-521 and E-cadherin, which both localize to stem cell niches in vivo. This matrix allows clonal derivation, clonal survival and long-term self-renewal of hES cells under completely chemically defined and xeno-free conditions without ROCK inhibitors. Neither LN-521 nor E-cadherin alone enable clonal survival of hES cells. The LN-521/E-cadherin matrix allows hES cell line derivation from blastocyst inner cell mass and single blastomere cells without a need to destroy the embryo. This method can facilitate the generation of hES cell lines for development of different cell types for regenerative medicine purposes.

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“…Most of the surface proteins will be destroyed by Trypsin detachment during the procedure. Trypsin needs to be actively inactivated to stop the reaction [16]. Similarly, the cytotoxic effect may also be present by the use of detergents [17,18].…”

Section: Discussionmentioning

confidence: 99%

Biocompatibility Issue Of Tissue Engineered Heart Valves

Wilczek¹

2015

Archives of Metallurgy and Materials

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Tissue engineering is a new field of knowledge which creates the possibilities for producing bioactive cardiac prostheses that will characterize by biomechanical and morphological properties similar to native tissue. It is expected that it will be characterized by high durability, which is very important from the social and clinical point of view. The aim of the study was to compare the cytotoxic effect of enzymatic and detergent acellularization methods commonly used for the biological scaffold preparation. It seems that the use of enzymatic methods, allows efficient donor cells removal while maintaining the ability to autologous cell seeding. Heart valves bioprosthesis created using these techniques, may be a good alternative to the currently used prostheses.Keywords: tissue engineering, heart valve, cytotoxicity, scaffold.Inżynieria tkankowa jest nową dziedziną wiedzy, która stwarza możliwości wytwarzania protez serca bioaktywnych charakteryzujących się właściwościami biomechanicznymi i morfologicznymi zbliżonymi do tkanki rodzimej. Oczekuje się, że proteza serca będzie charakteryzować się wysoką trwałością, co jest bardzo ważne z punktu widzenia społecznego i klinicznego. Celem badania było porównanie efektu cytotoksycznego enzymatycznych i detergentowych metod usuwania komórek, powszechnie stosowanych do wytwarzania biologicznych rusztowań. Wydaje się, że stosowanie metod enzymatycznych, umożliwia wydajne usunięcie komórek dawcy przy zachowaniu zdolności do autologicznego posiewu komórek. Bioprotezy zastawek serca tworzone za pomocą tej techniki mogą być dobrą alternatywą dla obecnie stosowanych mechanicznych i biologicznych protez zastawek serca.

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Efficient propagation of single cells accutase‐dissociated human embryonic stem cells

Cited by 145 publications

References 20 publications

High-efficiency siRNA-based gene knockdown in human embryonic stem cells

High-efficiency siRNA-based gene knockdown in human embryonic stem cells

Clonal culturing of human embryonic stem cells on laminin-521/E-cadherin matrix in defined and xeno-free environment

Biocompatibility Issue Of Tissue Engineered Heart Valves

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