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. 2017 May 22;8:103. doi: 10.3389/fendo.2017.00103

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Copyright © 2017 Arauchi, Matsuura, Shimizu and Okano.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Identification of thyroid transcription markers. (A) Pax8 and Nkx2-1 mRNA expression as measured with real-time reverse-transcription polymerase chain reaction. Levels of gene expression were normalized to Gapdh. Bars indicate average percentage of Gapdh gene expression ± SD (n = 3). ND indicates mRNA expression was not detected and NS indicates not significant. There was no significant difference between all samples in Pax8 by ANOVA. (B) Percentage of Pax8 and Nkx2-1 co-expressing cells as counted by flow cytometry. Pax8-positive cells increased in 5 days to more than 50%, which was maintained until day 20, except for a transient decrease on day 10. Percentage of Pax8 and Nkx2-1 double-positive cells increased in a time-dependent manner, resulting in 16% approximately on culture day 20. Negative controls were applied for normal IgG instead of primary antibodies. Bars indicate average percentage of Pax8- and Nkx2-1-coexpressing cells ± SD (n = 3). Statistical analysis was performed by one-way ANOVA. (C) Immunohistological analysis of Pax8 and Nkx2-1. Images in the upper line present negative controls. Images in lower show Pax8 (green), Nkx2-1 (red), and Hoechst33342 (blue). Scale bar = 50 µm.

Identification of thyroid transcription markers. (A) Pax8 and Nkx2-1 mRNA expression as measured with real-time reverse-transcription polymerase chain reaction. Levels of gene expression were normalized to Gapdh. Bars indicate average percentage of Gapdh gene expression ± SD (n = 3). ND indicates mRNA expression was not detected and NS indicates not significant. There was no significant difference between all samples in Pax8 by ANOVA. (B) Percentage of Pax8 and Nkx2-1 co-expressing cells as counted by flow cytometry. Pax8-positive cells increased in 5 days to more than 50%, which was maintained until day 20, except for a transient decrease on day 10. Percentage of Pax8 and Nkx2-1 double-positive cells increased in a time-dependent manner, resulting in 16% approximately on culture day 20. Negative controls were applied for normal IgG instead of primary antibodies. Bars indicate average percentage of Pax8- and Nkx2-1-coexpressing cells ± SD (n = 3). Statistical analysis was performed by one-way ANOVA. (C) Immunohistological analysis of Pax8 and Nkx2-1. Images in the upper line present negative controls. Images in lower show Pax8 (green), Nkx2-1 (red), and Hoechst33342 (blue). Scale bar = 50 µm.