Differentiating Human Embryonic Stem Cells into Neural Progenitors

In This Article

Abstract

Source: Jeon, K. et al., Efficient Neural Differentiation using Single-Cell Culture of Human Embryonic Stem Cells. J. Vis. Exp. (2020)

This video demonstrates the differentiation of human embryonic stem cells, or hESCs, directly into neural progenitor cells without using feeder cells or any intermediate steps. The selective inhibition of bone morphogenetic protein or BMP and transforming growth factor-beta, or TGF-β, signaling pathways leads to hESCs differentiation into neural progenitor cells, or NPCs.

Protocol

1. Preparation of human embryonic stem cells (hESC)-qualified Basement Membrane Matrix-coated Plates

Slowly thaw the hESC-qualified basement membrane matrix (see Table of Materials) solution at 4 °C for at least 2–3 h or overnight to avoid formation of a gel.
To prepare basement membrane matrix-coated plates, dilute matrix in cold Dulbecco's Modified Eagle Medium with F12 (DMEM+F12) to a 2% final concentration. Mix well and coat each well of a 6 well plate with 1 mL of the diluted matrix solution.
Incubate the basement membrane matrix-coated plates at room temperature (RT) for at least 3 h or at 4 °C ove....

Access restricted. Please log in or start a trial to view this content.

Results

Cell culture process diagram with growth chart and cell cycle analysis; microscopy images included.
Figure 1: Adaptation of colony type hESCs to single-cell type culture. (A) Representative phase contrast images of single cell cultures of H9 hESCs at different times after plating on 2% basement membrane matrix-coated dishes. Low (left).......