Simple Generation of a High Yield Culture of Induced Neurons from Human Adult Skin Fibroblasts
MyJove Corporation -- Journal of Visualized Experiments
DOI 10.3791/56904
Keyword(s)
  1. Neuroscience
  2. Issue 132
  3. Induced neurons
  4. adult human dermal fibroblasts
  5. direct neural reprogramming
  6. human neurons
  7. FACS purification
  8. lentiviral vector
Abstract(s)

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