A Novel Protocol for Detection of Senescence and Calcification Markers by Fluorescence Microscopy
Vascular calcification and stiffening of the arterial wall is a systemic phenomenon that is associated with aging and it can be increased by several risk factors. The underlying mechanisms, especially the pathways of cellular senescence, are under current investigation. Easily manageable in vitro se...
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| Format: | Article |
| Language: | English |
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MDPI AG
2020-05-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/21/10/3475 |
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| author | Jaqueline Herrmann Milen Babic Markus Tölle Kai-Uwe Eckardt Markus van der Giet Mirjam Schuchardt |
| author_facet | Jaqueline Herrmann Milen Babic Markus Tölle Kai-Uwe Eckardt Markus van der Giet Mirjam Schuchardt |
| author_sort | Jaqueline Herrmann |
| collection | DOAJ |
| description | Vascular calcification and stiffening of the arterial wall is a systemic phenomenon that is associated with aging and it can be increased by several risk factors. The underlying mechanisms, especially the pathways of cellular senescence, are under current investigation. Easily manageable in vitro settings help to study the signaling pathways. The experimental setting presented here is based on an in vitro model using rat vascular smooth muscle cells and the detection of senescence and osteoblastic markers via immunofluorescence and RNAscope™. Co-staining of the senescence marker p21, the osteoblastic marker osteopontin, detection of senescence-associated heterochromatin foci, and senescence-associated β-galactosidase is possible within one test approach requiring fewer cells. The protocol is a fast and reliable evaluation method for multiplexing of calcifying and senescence markers with fluorescence microscopy detection. The experimental setting enables analysis on single cell basis and allows detection of intra-individual variances of cultured cells. |
| first_indexed | 2024-03-10T19:49:31Z |
| format | Article |
| id | doaj.art-52a9b08d8603432eb7506af6a9b6ca09 |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-10T19:49:31Z |
| publishDate | 2020-05-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-52a9b08d8603432eb7506af6a9b6ca092023-11-20T00:29:45ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672020-05-012110347510.3390/ijms21103475A Novel Protocol for Detection of Senescence and Calcification Markers by Fluorescence MicroscopyJaqueline Herrmann0Milen Babic1Markus Tölle2Kai-Uwe Eckardt3Markus van der Giet4Mirjam Schuchardt5Department of Nephrology and Medical Intensive Care, Charité—Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universtität zu Berlin, and Berlin Institute of Health, Hindenburgdamm 30, 12203 Berlin, GermanyDepartment of Nephrology and Medical Intensive Care, Charité—Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universtität zu Berlin, and Berlin Institute of Health, Hindenburgdamm 30, 12203 Berlin, GermanyDepartment of Nephrology and Medical Intensive Care, Charité—Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universtität zu Berlin, and Berlin Institute of Health, Hindenburgdamm 30, 12203 Berlin, GermanyDepartment of Nephrology and Medical Intensive Care, Charité—Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universtität zu Berlin, and Berlin Institute of Health, Hindenburgdamm 30, 12203 Berlin, GermanyDepartment of Nephrology and Medical Intensive Care, Charité—Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universtität zu Berlin, and Berlin Institute of Health, Hindenburgdamm 30, 12203 Berlin, GermanyDepartment of Nephrology and Medical Intensive Care, Charité—Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universtität zu Berlin, and Berlin Institute of Health, Hindenburgdamm 30, 12203 Berlin, GermanyVascular calcification and stiffening of the arterial wall is a systemic phenomenon that is associated with aging and it can be increased by several risk factors. The underlying mechanisms, especially the pathways of cellular senescence, are under current investigation. Easily manageable in vitro settings help to study the signaling pathways. The experimental setting presented here is based on an in vitro model using rat vascular smooth muscle cells and the detection of senescence and osteoblastic markers via immunofluorescence and RNAscope™. Co-staining of the senescence marker p21, the osteoblastic marker osteopontin, detection of senescence-associated heterochromatin foci, and senescence-associated β-galactosidase is possible within one test approach requiring fewer cells. The protocol is a fast and reliable evaluation method for multiplexing of calcifying and senescence markers with fluorescence microscopy detection. The experimental setting enables analysis on single cell basis and allows detection of intra-individual variances of cultured cells.https://www.mdpi.com/1422-0067/21/10/3475calcificationsenescencesmooth muscle cellSA-β-galactosidasesenescence-associated heterochromatin foci |
| spellingShingle | Jaqueline Herrmann Milen Babic Markus Tölle Kai-Uwe Eckardt Markus van der Giet Mirjam Schuchardt A Novel Protocol for Detection of Senescence and Calcification Markers by Fluorescence Microscopy International Journal of Molecular Sciences calcification senescence smooth muscle cell SA-β-galactosidase senescence-associated heterochromatin foci |
| title | A Novel Protocol for Detection of Senescence and Calcification Markers by Fluorescence Microscopy |
| title_full | A Novel Protocol for Detection of Senescence and Calcification Markers by Fluorescence Microscopy |
| title_fullStr | A Novel Protocol for Detection of Senescence and Calcification Markers by Fluorescence Microscopy |
| title_full_unstemmed | A Novel Protocol for Detection of Senescence and Calcification Markers by Fluorescence Microscopy |
| title_short | A Novel Protocol for Detection of Senescence and Calcification Markers by Fluorescence Microscopy |
| title_sort | novel protocol for detection of senescence and calcification markers by fluorescence microscopy |
| topic | calcification senescence smooth muscle cell SA-β-galactosidase senescence-associated heterochromatin foci |
| url | https://www.mdpi.com/1422-0067/21/10/3475 |
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