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. 2020 Aug 12;10:13605. doi: 10.1038/s41598-020-70707-6

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© The Author(s) 2020

Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Effects of nano-warming on hiPSC cryopreservation. (a) Schematic illustration for cryopreservation of iPSC cells. hiPSCs were applied to a vial at 1 × 10⁶ cells/mL in StemCell Keep containing magnetite nanoparticles at 5 mg/mL. The vial was directly immersed in liquid nitrogen for 15 min. The vial was then irradiated with an alternating magnetic field at 10 kW, 208 kHz. (b) Effects of nano-warming on hiPSC viability. After freezing and thawing, cell viability was assayed using a cell viability imaging kit based on Hoechst 33342 for live cells and SYTOX green nucleic acid stain for dead cells. Open columns, convective warming (water bath at 37 °C) of magnetite-loaded StemCell Keep; Closed columns, nano-warming of magnetite-loaded StemCell Keep. Data are expressed as the mean ± SD (n = 3). *P < 0.05. (c) Alkaline phosphatase (AP) staining of hiPSC colonies after nano-warming. Control, non-frozen hiPSCs. Data are expressed as the mean ± SD (n = 3). (d) RT-PCR analyses of lineage-specific genes. After nano-warming, hiPSCs were cultured under each differentiation condition for the three germ layers: endoderm, mesoderm, and ectoderm. C non-frozen control, N nano-warming. Relative expression levels normalized to β-actin gene expression are shown below the bands.