Simple Generation of a High Yield Culture of Induced Neurons from Human Adult Skin Fibroblasts

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Simple Generation of a High Yield Culture of Induced Neurons from Human Adult Skin Fibroblasts. / Shrigley, Shelby; Pircs, Karolina; Barker, Roger A.; Parmar, Malin; Drouin-ouellet, Janelle.

I: Journal of Visualized Experiments, Vol. 132, e56904, 05.02.2018.

Forskningsoutput: TidskriftsbidragArtikel i vetenskaplig tidskrift

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T1 - Simple Generation of a High Yield Culture of Induced Neurons from Human Adult Skin Fibroblasts

AU - Shrigley, Shelby

AU - Pircs, Karolina

AU - Barker, Roger A.

AU - Parmar, Malin

AU - Drouin-ouellet, Janelle

PY - 2018/2/5

Y1 - 2018/2/5

N2 - Induced neurons (iNs), the product of somatic cells directly converted to neurons, are a way to obtain patient-derived neurons from tissue thatis easily accessible. Through this route, mature neurons can be obtained in a matter of a few weeks. Here, we describe a straightforward andrapid one-step protocol to obtain iNs from dermal fibroblasts obtained through biopsy samples from adult human donors. We explain each stepof the process, including the maintenance of the dermal fibroblasts, the freezing procedure to build a stock of the cell line, seeding of the cellsfor reprogramming, as well as the culture conditions during the conversion process. In addition, we describe the preparation of glass coverslipsfor electrophysiological recordings, long-term coating conditions, and fluorescence activated cell sorting (FACS). We also illustrate examplesof the results to be expected. The protocol described here is easy to perform and can be applied to human fibroblasts derived from human skinbiopsies from patients with various different diagnoses and ages. This protocol generates a sufficient amount of iNs which can be used for a widearray of biomedical applications, including disease modeling, drug screening, and target validation.

AB - Induced neurons (iNs), the product of somatic cells directly converted to neurons, are a way to obtain patient-derived neurons from tissue thatis easily accessible. Through this route, mature neurons can be obtained in a matter of a few weeks. Here, we describe a straightforward andrapid one-step protocol to obtain iNs from dermal fibroblasts obtained through biopsy samples from adult human donors. We explain each stepof the process, including the maintenance of the dermal fibroblasts, the freezing procedure to build a stock of the cell line, seeding of the cellsfor reprogramming, as well as the culture conditions during the conversion process. In addition, we describe the preparation of glass coverslipsfor electrophysiological recordings, long-term coating conditions, and fluorescence activated cell sorting (FACS). We also illustrate examplesof the results to be expected. The protocol described here is easy to perform and can be applied to human fibroblasts derived from human skinbiopsies from patients with various different diagnoses and ages. This protocol generates a sufficient amount of iNs which can be used for a widearray of biomedical applications, including disease modeling, drug screening, and target validation.

U2 - 10.3791/56904

DO - 10.3791/56904

M3 - Article

C2 - 29443113

VL - 132

JO - Journal of Visualized Experiments

JF - Journal of Visualized Experiments

SN - 1940-087X

M1 - e56904

ER -