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A) Time-lapse images of a live plated neurocrestoid with nuclei stained with SPY650-DNA. B) Representative fluorescence images at day 14 of plated neurocrestoids immunostained for SOX10 and nuclei (DAPI). C) Bar chart depicting percentage of SOX10 positive cells amongst the total nuclei (DAPI) in plated neurocrestoids at day 14 across experiments from 3 independent hPSC lines (mean ± S.D). Each point represents the average across n=5-18 organoids per individual experiment. F) Neurocrestoids plated at day 14 and stained against HOXC9, SOX10 and nuclei (DAPI). E) Representative immunofluorescence images of mouse trunk explant and human neurocrestoid-derived NCCs stained for SOX10. F) Bar charts indicating the area, circularity and solidity values for neurocrestoid-derived human and mouse trunk NC nuclei (mean ± S.D). Points represent the average value per field of view. For the human datasets, a total of 38 datapoints are plotted representing the average value per field of view, 5 images per sample, 3-5 organoids per cell line from 3 <t>hiPSC</t> <t>lines</t> (Control 3, KOLFC21 and KOLF2.1J) in 3 experimental blocks. The mouse dataset consists of the average of 10 cells per image, 10 images per explant in 3 mouse trunk NC explants cultured for 48 to 72 hours in 2 experimental blocks. Non-parametric Mann-Whitney test to test for significance between conditions, **P<0.01, ns: non-significant.
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A) Time-lapse images of a live plated neurocrestoid with nuclei stained with SPY650-DNA. B) Representative fluorescence images at day 14 of plated neurocrestoids immunostained for SOX10 and nuclei (DAPI). C) Bar chart depicting percentage of SOX10 positive cells amongst the total nuclei (DAPI) in plated neurocrestoids at day 14 across experiments from 3 independent hPSC lines (mean ± S.D). Each point represents the average across n=5-18 organoids per individual experiment. F) Neurocrestoids plated at day 14 and stained against HOXC9, SOX10 and nuclei (DAPI). E) Representative immunofluorescence images of mouse trunk explant and human neurocrestoid-derived NCCs stained for SOX10. F) Bar charts indicating the area, circularity and solidity values for neurocrestoid-derived human and mouse trunk NC nuclei (mean ± S.D). Points represent the average value per field of view. For the human datasets, a total of 38 datapoints are plotted representing the average value per field of view, 5 images per sample, 3-5 organoids per cell line from 3 <t>hiPSC</t> <t>lines</t> (Control 3, KOLFC21 and KOLF2.1J) in 3 experimental blocks. The mouse dataset consists of the average of 10 cells per image, 10 images per explant in 3 mouse trunk NC explants cultured for 48 to 72 hours in 2 experimental blocks. Non-parametric Mann-Whitney test to test for significance between conditions, **P<0.01, ns: non-significant.
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A) Time-lapse images of a live plated neurocrestoid with nuclei stained with SPY650-DNA. B) Representative fluorescence images at day 14 of plated neurocrestoids immunostained for SOX10 and nuclei (DAPI). C) Bar chart depicting percentage of SOX10 positive cells amongst the total nuclei (DAPI) in plated neurocrestoids at day 14 across experiments from 3 independent hPSC lines (mean ± S.D). Each point represents the average across n=5-18 organoids per individual experiment. F) Neurocrestoids plated at day 14 and stained against HOXC9, SOX10 and nuclei (DAPI). E) Representative immunofluorescence images of mouse trunk explant and human neurocrestoid-derived NCCs stained for SOX10. F) Bar charts indicating the area, circularity and solidity values for neurocrestoid-derived human and mouse trunk NC nuclei (mean ± S.D). Points represent the average value per field of view. For the human datasets, a total of 38 datapoints are plotted representing the average value per field of view, 5 images per sample, 3-5 organoids per cell line from 3 <t>hiPSC</t> <t>lines</t> (Control 3, KOLFC21 and KOLF2.1J) in 3 experimental blocks. The mouse dataset consists of the average of 10 cells per image, 10 images per explant in 3 mouse trunk NC explants cultured for 48 to 72 hours in 2 experimental blocks. Non-parametric Mann-Whitney test to test for significance between conditions, **P<0.01, ns: non-significant.
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A) Time-lapse images of a live plated neurocrestoid with nuclei stained with SPY650-DNA. B) Representative fluorescence images at day 14 of plated neurocrestoids immunostained for SOX10 and nuclei (DAPI). C) Bar chart depicting percentage of SOX10 positive cells amongst the total nuclei (DAPI) in plated neurocrestoids at day 14 across experiments from 3 independent hPSC lines (mean ± S.D). Each point represents the average across n=5-18 organoids per individual experiment. F) Neurocrestoids plated at day 14 and stained against HOXC9, SOX10 and nuclei (DAPI). E) Representative immunofluorescence images of mouse trunk explant and human neurocrestoid-derived NCCs stained for SOX10. F) Bar charts indicating the area, circularity and solidity values for neurocrestoid-derived human and mouse trunk NC nuclei (mean ± S.D). Points represent the average value per field of view. For the human datasets, a total of 38 datapoints are plotted representing the average value per field of view, 5 images per sample, 3-5 organoids per cell line from 3 <t>hiPSC</t> <t>lines</t> (Control 3, KOLFC21 and KOLF2.1J) in 3 experimental blocks. The mouse dataset consists of the average of 10 cells per image, 10 images per explant in 3 mouse trunk NC explants cultured for 48 to 72 hours in 2 experimental blocks. Non-parametric Mann-Whitney test to test for significance between conditions, **P<0.01, ns: non-significant.
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A) Time-lapse images of a live plated neurocrestoid with nuclei stained with SPY650-DNA. B) Representative fluorescence images at day 14 of plated neurocrestoids immunostained for SOX10 and nuclei (DAPI). C) Bar chart depicting percentage of SOX10 positive cells amongst the total nuclei (DAPI) in plated neurocrestoids at day 14 across experiments from 3 independent hPSC lines (mean ± S.D). Each point represents the average across n=5-18 organoids per individual experiment. F) Neurocrestoids plated at day 14 and stained against HOXC9, SOX10 and nuclei (DAPI). E) Representative immunofluorescence images of mouse trunk explant and human neurocrestoid-derived NCCs stained for SOX10. F) Bar charts indicating the area, circularity and solidity values for neurocrestoid-derived human and mouse trunk NC nuclei (mean ± S.D). Points represent the average value per field of view. For the human datasets, a total of 38 datapoints are plotted representing the average value per field of view, 5 images per sample, 3-5 organoids per cell line from 3 <t>hiPSC</t> <t>lines</t> (Control 3, KOLFC21 and KOLF2.1J) in 3 experimental blocks. The mouse dataset consists of the average of 10 cells per image, 10 images per explant in 3 mouse trunk NC explants cultured for 48 to 72 hours in 2 experimental blocks. Non-parametric Mann-Whitney test to test for significance between conditions, **P<0.01, ns: non-significant.
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A) Time-lapse images of a live plated neurocrestoid with nuclei stained with SPY650-DNA. B) Representative fluorescence images at day 14 of plated neurocrestoids immunostained for SOX10 and nuclei (DAPI). C) Bar chart depicting percentage of SOX10 positive cells amongst the total nuclei (DAPI) in plated neurocrestoids at day 14 across experiments from 3 independent hPSC lines (mean ± S.D). Each point represents the average across n=5-18 organoids per individual experiment. F) Neurocrestoids plated at day 14 and stained against HOXC9, SOX10 and nuclei (DAPI). E) Representative immunofluorescence images of mouse trunk explant and human neurocrestoid-derived NCCs stained for SOX10. F) Bar charts indicating the area, circularity and solidity values for neurocrestoid-derived human and mouse trunk NC nuclei (mean ± S.D). Points represent the average value per field of view. For the human datasets, a total of 38 datapoints are plotted representing the average value per field of view, 5 images per sample, 3-5 organoids per cell line from 3 <t>hiPSC</t> <t>lines</t> (Control 3, KOLFC21 and KOLF2.1J) in 3 experimental blocks. The mouse dataset consists of the average of 10 cells per image, 10 images per explant in 3 mouse trunk NC explants cultured for 48 to 72 hours in 2 experimental blocks. Non-parametric Mann-Whitney test to test for significance between conditions, **P<0.01, ns: non-significant.
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A) Time-lapse images of a live plated neurocrestoid with nuclei stained with SPY650-DNA. B) Representative fluorescence images at day 14 of plated neurocrestoids immunostained for SOX10 and nuclei (DAPI). C) Bar chart depicting percentage of SOX10 positive cells amongst the total nuclei (DAPI) in plated neurocrestoids at day 14 across experiments from 3 independent hPSC lines (mean ± S.D). Each point represents the average across n=5-18 organoids per individual experiment. F) Neurocrestoids plated at day 14 and stained against HOXC9, SOX10 and nuclei (DAPI). E) Representative immunofluorescence images of mouse trunk explant and human neurocrestoid-derived NCCs stained for SOX10. F) Bar charts indicating the area, circularity and solidity values for neurocrestoid-derived human and mouse trunk NC nuclei (mean ± S.D). Points represent the average value per field of view. For the human datasets, a total of 38 datapoints are plotted representing the average value per field of view, 5 images per sample, 3-5 organoids per cell line from 3 hiPSC lines (Control 3, KOLFC21 and KOLF2.1J) in 3 experimental blocks. The mouse dataset consists of the average of 10 cells per image, 10 images per explant in 3 mouse trunk NC explants cultured for 48 to 72 hours in 2 experimental blocks. Non-parametric Mann-Whitney test to test for significance between conditions, **P<0.01, ns: non-significant.

Journal: bioRxiv

Article Title: Dynamic modelling of human neural crest development using a bioengineered stem cell organoid system

doi: 10.64898/2026.05.04.721958

Figure Lengend Snippet: A) Time-lapse images of a live plated neurocrestoid with nuclei stained with SPY650-DNA. B) Representative fluorescence images at day 14 of plated neurocrestoids immunostained for SOX10 and nuclei (DAPI). C) Bar chart depicting percentage of SOX10 positive cells amongst the total nuclei (DAPI) in plated neurocrestoids at day 14 across experiments from 3 independent hPSC lines (mean ± S.D). Each point represents the average across n=5-18 organoids per individual experiment. F) Neurocrestoids plated at day 14 and stained against HOXC9, SOX10 and nuclei (DAPI). E) Representative immunofluorescence images of mouse trunk explant and human neurocrestoid-derived NCCs stained for SOX10. F) Bar charts indicating the area, circularity and solidity values for neurocrestoid-derived human and mouse trunk NC nuclei (mean ± S.D). Points represent the average value per field of view. For the human datasets, a total of 38 datapoints are plotted representing the average value per field of view, 5 images per sample, 3-5 organoids per cell line from 3 hiPSC lines (Control 3, KOLFC21 and KOLF2.1J) in 3 experimental blocks. The mouse dataset consists of the average of 10 cells per image, 10 images per explant in 3 mouse trunk NC explants cultured for 48 to 72 hours in 2 experimental blocks. Non-parametric Mann-Whitney test to test for significance between conditions, **P<0.01, ns: non-significant.

Article Snippet: HiPSC lines were obtained either from commercially available sources: Control 3 (ThermoFisher Scientific, A18945), KOLF2.1J (Jackson Laboratory, JIPSC1000), or kindly donated: KOLF2C1 (Maximiliano Gutierrez, The Francis Crick Institute).

Techniques: Staining, Fluorescence, Immunofluorescence, Derivative Assay, Control, Cell Culture, MANN-WHITNEY