Drying of Functional Hydrogels: Development of a Workflow for Bioreactor-Integrated Freeze-Drying of Protein-Coated Alginate Microcarriers for iPS Cell-Based Screenings
Protein-coated ultra-high viscosity (UHV)-alginate hydrogels are essential to mimic the physiological in vivo environment of humans in several in vitro applications. This work presents an optimized bioreactor-integrated freeze-drying process for Matrigel<sup>TM</sup>-coated UHV-alginate...
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MDPI AG
2025-06-01
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| Online Access: | https://www.mdpi.com/2310-2861/11/6/439 |
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| author | Johnn Majd Balsters Alexander Bäumchen Michael Roland Stefan Diebels Julia C. Neubauer Michael M. Gepp Heiko Zimmermann |
| author_facet | Johnn Majd Balsters Alexander Bäumchen Michael Roland Stefan Diebels Julia C. Neubauer Michael M. Gepp Heiko Zimmermann |
| author_sort | Johnn Majd Balsters |
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| description | Protein-coated ultra-high viscosity (UHV)-alginate hydrogels are essential to mimic the physiological in vivo environment of humans in several in vitro applications. This work presents an optimized bioreactor-integrated freeze-drying process for Matrigel<sup>TM</sup>-coated UHV-alginate microcarriers in the context of human induced pluripotent stem cell (hiPSC) expansion. The impact of freeze-drying on the UHV-alginate microcarriers using trehalose 100 mg/mL in 0.9% NaCl as a lyoprotective agent, as well as the stem cell response using hiPSCs, was analyzed using microscopy-based screenings. First observations of the process showed that the integrity of the cake was preserved in the samples with a maximum vapor exchanging rate. Following rehydration, the UHV-alginate microcarriers retained their original morphology. Upon the addition of Poloxamer 188, stickiness and bubble formation were reduced. The expansion of hiPSCs in a suspension bioreactor resulted in a 5–7-fold increase in total cell count, yielding at least 1.3 × 10<sup>7</sup> cells with viability exceeding 80% after seven days of cultivation. In flow cytometry analysis, the pluripotency factors OCT3/4 and SSEA4 resulted in positive signals in over 98% of cells, while the differentiation factor SSEA1 was positive in fewer than 10% of cells. Supported by preceding in silico predictions of drying time, this study presents, for the first time, basic steps toward a “ready-to-use” bioreactor-integrated freeze-drying process for UHV-alginate microcarriers in the iPSC context. |
| format | Article |
| id | doaj-art-d6f7e9f324de48be9df0bb9fdbf4e8c5 |
| institution | Matheson Library |
| issn | 2310-2861 |
| language | English |
| publishDate | 2025-06-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Gels |
| spelling | doaj-art-d6f7e9f324de48be9df0bb9fdbf4e8c52025-06-25T13:53:51ZengMDPI AGGels2310-28612025-06-0111643910.3390/gels11060439Drying of Functional Hydrogels: Development of a Workflow for Bioreactor-Integrated Freeze-Drying of Protein-Coated Alginate Microcarriers for iPS Cell-Based ScreeningsJohnn Majd Balsters0Alexander Bäumchen1Michael Roland2Stefan Diebels3Julia C. Neubauer4Michael M. Gepp5Heiko Zimmermann6Fraunhofer Institute for Biomedical Engineering IBMT, 66280 Sulzbach, GermanyApplied Mechanics, Saarland University, Campus, 66123 Saarbruecken, GermanyApplied Mechanics, Saarland University, Campus, 66123 Saarbruecken, GermanyApplied Mechanics, Saarland University, Campus, 66123 Saarbruecken, GermanyFraunhofer Institute for Biomedical Engineering IBMT, 66280 Sulzbach, GermanyFraunhofer Institute for Biomedical Engineering IBMT, 66280 Sulzbach, GermanyFraunhofer Institute for Biomedical Engineering IBMT, 66280 Sulzbach, GermanyProtein-coated ultra-high viscosity (UHV)-alginate hydrogels are essential to mimic the physiological in vivo environment of humans in several in vitro applications. This work presents an optimized bioreactor-integrated freeze-drying process for Matrigel<sup>TM</sup>-coated UHV-alginate microcarriers in the context of human induced pluripotent stem cell (hiPSC) expansion. The impact of freeze-drying on the UHV-alginate microcarriers using trehalose 100 mg/mL in 0.9% NaCl as a lyoprotective agent, as well as the stem cell response using hiPSCs, was analyzed using microscopy-based screenings. First observations of the process showed that the integrity of the cake was preserved in the samples with a maximum vapor exchanging rate. Following rehydration, the UHV-alginate microcarriers retained their original morphology. Upon the addition of Poloxamer 188, stickiness and bubble formation were reduced. The expansion of hiPSCs in a suspension bioreactor resulted in a 5–7-fold increase in total cell count, yielding at least 1.3 × 10<sup>7</sup> cells with viability exceeding 80% after seven days of cultivation. In flow cytometry analysis, the pluripotency factors OCT3/4 and SSEA4 resulted in positive signals in over 98% of cells, while the differentiation factor SSEA1 was positive in fewer than 10% of cells. Supported by preceding in silico predictions of drying time, this study presents, for the first time, basic steps toward a “ready-to-use” bioreactor-integrated freeze-drying process for UHV-alginate microcarriers in the iPSC context.https://www.mdpi.com/2310-2861/11/6/439suspension bioreactordrug discoverypluripotent stem cellsfreeze-drying(UHV)-alginatetissue engineering |
| spellingShingle | Johnn Majd Balsters Alexander Bäumchen Michael Roland Stefan Diebels Julia C. Neubauer Michael M. Gepp Heiko Zimmermann Drying of Functional Hydrogels: Development of a Workflow for Bioreactor-Integrated Freeze-Drying of Protein-Coated Alginate Microcarriers for iPS Cell-Based Screenings Gels suspension bioreactor drug discovery pluripotent stem cells freeze-drying (UHV)-alginate tissue engineering |
| title | Drying of Functional Hydrogels: Development of a Workflow for Bioreactor-Integrated Freeze-Drying of Protein-Coated Alginate Microcarriers for iPS Cell-Based Screenings |
| title_full | Drying of Functional Hydrogels: Development of a Workflow for Bioreactor-Integrated Freeze-Drying of Protein-Coated Alginate Microcarriers for iPS Cell-Based Screenings |
| title_fullStr | Drying of Functional Hydrogels: Development of a Workflow for Bioreactor-Integrated Freeze-Drying of Protein-Coated Alginate Microcarriers for iPS Cell-Based Screenings |
| title_full_unstemmed | Drying of Functional Hydrogels: Development of a Workflow for Bioreactor-Integrated Freeze-Drying of Protein-Coated Alginate Microcarriers for iPS Cell-Based Screenings |
| title_short | Drying of Functional Hydrogels: Development of a Workflow for Bioreactor-Integrated Freeze-Drying of Protein-Coated Alginate Microcarriers for iPS Cell-Based Screenings |
| title_sort | drying of functional hydrogels development of a workflow for bioreactor integrated freeze drying of protein coated alginate microcarriers for ips cell based screenings |
| topic | suspension bioreactor drug discovery pluripotent stem cells freeze-drying (UHV)-alginate tissue engineering |
| url | https://www.mdpi.com/2310-2861/11/6/439 |
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