Comparison of 2D and 3D neural induction methods for the generation of neural progenitor cells from human induced pluripotent stem cells

Abinaya Chandrasekaran, Hasan X. Avci, Anna Ochalek, Lone N. Rösingh, Kinga Molnár, L. László, Tamás Bellák, Annamária Téglási, Krisztina Pesti, A. Mike, Phetcharat Phanthong, Orsolya Bíró, Vanessa Hall, Narisorn Kitiyanant, Karl Heinz Krause, Julianna Kobolák, A. Dinnyés

Research output: Contribution to journalArticle

20 Citations (Scopus)


Neural progenitor cells (NPCs) from human induced pluripotent stem cells (hiPSCs) are frequently induced using 3D culture methodologies however, it is unknown whether spheroid-based (3D) neural induction is actually superior to monolayer (2D) neural induction. Our aim was to compare the efficiency of 2D induction with 3D induction method in their ability to generate NPCs, and subsequently neurons and astrocytes. Neural differentiation was analysed at the protein level qualitatively by immunocytochemistry and quantitatively by flow cytometry for NPC (SOX1, PAX6, NESTIN), neuronal (MAP2, TUBB3), cortical layer (TBR1, CUX1) and glial markers (SOX9, GFAP, AQP4). Electron microscopy demonstrated that both methods resulted in morphologically similar neural rosettes. However, quantification of NPCs derived from 3D neural induction exhibited an increase in the number of PAX6/NESTIN double positive cells and the derived neurons exhibited longer neurites. In contrast, 2D neural induction resulted in more SOX1 positive cells. While 2D monolayer induction resulted in slightly less mature neurons, at an early stage of differentiation, the patch clamp analysis failed to reveal any significant differences between the electrophysiological properties between the two induction methods. In conclusion, 3D neural induction increases the yield of PAX6+/NESTIN+ cells and gives rise to neurons with longer neurites, which might be an advantage for the production of forebrain cortical neurons, highlighting the potential of 3D neural induction, independent of iPSCs′ genetic background.

Original languageEnglish
Pages (from-to)139-151
Number of pages13
JournalStem Cell Research
Publication statusPublished - Dec 1 2017



  • 2D-3D neural induction
  • Electron microscopy
  • hiPSC
  • Neural induction
  • Neural progenitor cells
  • Patch clamp

ASJC Scopus subject areas

  • Developmental Biology
  • Cell Biology

Cite this

Chandrasekaran, A., Avci, H. X., Ochalek, A., Rösingh, L. N., Molnár, K., László, L., Bellák, T., Téglási, A., Pesti, K., Mike, A., Phanthong, P., Bíró, O., Hall, V., Kitiyanant, N., Krause, K. H., Kobolák, J., & Dinnyés, A. (2017). Comparison of 2D and 3D neural induction methods for the generation of neural progenitor cells from human induced pluripotent stem cells. Stem Cell Research, 25, 139-151.