Histological processing of un-/cellularized thermosensitive electrospun scaffolds

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dc.identifier.uri http://dx.doi.org/10.15488/4296
dc.identifier.uri https://www.repo.uni-hannover.de/handle/123456789/4330
dc.contributor.author Fuchs, Julia
dc.contributor.author Mueller, Marc
dc.contributor.author Daxböck, Christine
dc.contributor.author Stückler, Manuela
dc.contributor.author Lang, Ingrid
dc.contributor.author Leitinger, Gerd
dc.contributor.author Bock, Elisabeth
dc.contributor.author El-Heliebi, Amin
dc.contributor.author Moser, Gerit
dc.contributor.author Glasmacher, Birgit
dc.contributor.author Brislinger, Dagmar
dc.date.accessioned 2019-01-11T08:57:40Z
dc.date.available 2019-01-11T08:57:40Z
dc.date.issued 2018
dc.identifier.citation Fuchs, J.; Mueller, M.; Daxböck, C.; Stückler, M.; Lang, I. et al.: Histological processing of un-/cellularized thermosensitive electrospun scaffolds. In: Histochemistry and Cell Biology 18 (2018), Nr. 12, 4247. DOI: https://doi.org/10.1007/s00418-018-1757-7
dc.description.abstract Histological processing of thermosensitive electrospun poly(ε-caprolactone)/poly(l-lactide) (PCL/PLA) scaffolds fails, as poly(ε-caprolactone) (PCL) is characterized by its low-melting temperature (Tm = 60 °C). Here, we present an optimized low-temperature preparation method for the histological processing of un-/cellularized thermosensitive PCL/PLA scaffolds. Our study is aimed at the establishment of an optimized dehydration and low-melting-point paraffin-embedding method of electrospun PCL/PLA scaffolds (un-/cellularized). Furthermore, we compared this method with (a) automatized dehydration and standard paraffin embedding, (b) gelatin embedding followed by automatized dehydration and standard paraffin embedding, (c) cryofixation, and (d) acrylic resin embedding methods. We investigated pepsin and proteinase K antigen retrieval for their efficiency in epitope demasking at low temperatures and evaluated protocols for immunohistochemistry and immunofluorescence for cytokeratin 7 (CK7) and in situ padlock probe technology for beta actin (ACTB). Optimized dehydration and low-melting-point paraffin embedding preserved the PCL/PLA scaffold, as the diameter and structure of its fibers were unchanged. Cells attached to the PCL/PLA scaffolds showed limited alterations in size and morphology compared to control. Epitope demasking by enzymatic pepsin digestion and immunostaining of CK7 displayed an invasion of attached cells into the scaffold. Expression of ACTB and CK7 was shown by a combination of mRNA-based in situ padlock probe technology and immunofluorescence. In contrast, gelatin stabilization followed by standard paraffin embedding led to an overall shrinkage and melting of fibers, and therefore, no further analysis was possible. Acrylic resin embedding and cyrofixation caused fiber structures that were nearly unchanged in size and diameter. However, acrylic resin-embedded scaffolds are limited to 3 µm sections, whereas cyrofixation led to a reduction of the cell size by 14% compared to low-melting paraffin embedding. The combination of low-melting-point paraffin embedding and pepsin digestion as an antigen retrieval method offers a successful opportunity for histological investigations in thermosensitive specimens. eng
dc.language.iso eng
dc.publisher Heidelberg : Springer Verlag
dc.relation.ispartofseries Histochemistry and Cell Biology 18 (2018), Nr. 12
dc.rights CC BY 4.0 Unported
dc.rights.uri https://creativecommons.org/licenses/by/4.0/
dc.subject Cellularized prosthesis eng
dc.subject Graft eng
dc.subject Paraffin embedding eng
dc.subject PCL eng
dc.subject Polycaprolactone eng
dc.subject Polylactide acid eng
dc.subject.ddc 610 | Medizin, Gesundheit ger
dc.title Histological processing of un-/cellularized thermosensitive electrospun scaffolds
dc.type article
dc.type Text
dc.relation.issn 0948-6143
dc.relation.doi https://doi.org/10.1007/s00418-018-1757-7
dc.bibliographicCitation.issue 12
dc.bibliographicCitation.volume 18
dc.bibliographicCitation.firstPage 4247
dc.description.version publishedVersion
tib.accessRights frei zug�nglich

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