A multi-color flow cytometric method for characterizing murine reticulated platelets using SYTO 13
Reticulated platelets (RP) are immature platelets with heightened RNA content. An increased level of RP in the circulation is associated with various pathological conditions. In this study, we employed a novel flow cytometry approach for RP detection in mice, utilizing the nucleic acid dye SYTO 13 i...
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| Format: | Article |
| Language: | English |
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Taylor & Francis Group
2025-12-01
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| Series: | Platelets |
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| Online Access: | https://www.tandfonline.com/doi/10.1080/09537104.2025.2489020 |
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| author | Muataz Ali Hamad Krystin Krauel Nancy Schanze Mark Zurek Christoph B. Olivier Peter Kohl Thomas G. Nührenberg Daniel Duerschmied |
| author_facet | Muataz Ali Hamad Krystin Krauel Nancy Schanze Mark Zurek Christoph B. Olivier Peter Kohl Thomas G. Nührenberg Daniel Duerschmied |
| author_sort | Muataz Ali Hamad |
| collection | DOAJ |
| description | Reticulated platelets (RP) are immature platelets with heightened RNA content. An increased level of RP in the circulation is associated with various pathological conditions. In this study, we employed a novel flow cytometry approach for RP detection in mice, utilizing the nucleic acid dye SYTO 13 in conjunction with a platelet-specific marker (anti-mouse CD42b-DyLight 649). The efficacy of SYTO 13 for RP identification was confirmed by higher circulating RP levels in platelet-depleted mice during the recovery phase (35% ± 13%) compared to untreated mice (11% ± 1%, n = 9, p < .0001). We further characterized murine RP by exploring the surface expression of the platelet activation marker P-selectin. While there was no preactivation at baseline, stimulation with a thrombin receptor agonist (PAR-4) resulted in a higher increase in the percentage of P-selectin-positive platelets in RP (93% ± 6%) compared to non-RP (77% ± 14%, n = 6, p = .0065). In addition, RP exhibited higher geometric mean fluorescence intensity levels than non-RP (1874 ± 1278 versus 859 ± 549, n = 6, p = .02). Our proof-of-principle study demonstrates the efficacy of SYTO 13 in combination with platelet (activation) markers for identifying RP in both resting and activated states. This method holds promise for simultaneously monitoring RP levels and platelet activation in murine models of human disease. |
| format | Article |
| id | doaj-art-e7a0ab0c9af54e4da3d64670b1b828a0 |
| institution | Kabale University |
| issn | 0953-7104 1369-1635 |
| language | English |
| publishDate | 2025-12-01 |
| publisher | Taylor & Francis Group |
| record_format | Article |
| series | Platelets |
| spelling | doaj-art-e7a0ab0c9af54e4da3d64670b1b828a02025-08-20T03:53:22ZengTaylor & Francis GroupPlatelets0953-71041369-16352025-12-0136110.1080/09537104.2025.2489020A multi-color flow cytometric method for characterizing murine reticulated platelets using SYTO 13Muataz Ali Hamad0Krystin Krauel1Nancy Schanze2Mark Zurek3Christoph B. Olivier4Peter Kohl5Thomas G. Nührenberg6Daniel Duerschmied7Department of Cardiology and Angiology, Heart Center, Faculty of Medicine, University of Freiburg, Freiburg im Breisgau, GermanyDepartment of Cardiology and Angiology, Heart Center, Faculty of Medicine, University of Freiburg, Freiburg im Breisgau, GermanyDepartment of Cardiology and Angiology, Heart Center, Faculty of Medicine, University of Freiburg, Freiburg im Breisgau, GermanyDepartment of Cardiology and Angiology, Heart Center, Faculty of Medicine, University of Freiburg, Freiburg im Breisgau, GermanyDepartment of Cardiology and Angiology, Heart Center, Faculty of Medicine, University of Freiburg, Freiburg im Breisgau, GermanyInstitute for Experimental Cardiovascular Medicine, Heart Center, Faculty of Medicine, University of Freiburg, Freiburg im Breisgau, GermanyDepartment of Cardiology and Angiology, Heart Center, Faculty of Medicine, University of Freiburg, Freiburg im Breisgau, GermanyDepartment of Cardiology, Angiology, Haemostaseology, and Medical Intensive Care, Medical Centre Mannheim, Medical Faculty Mannheim, Heidelberg University, Mannheim, GermanyReticulated platelets (RP) are immature platelets with heightened RNA content. An increased level of RP in the circulation is associated with various pathological conditions. In this study, we employed a novel flow cytometry approach for RP detection in mice, utilizing the nucleic acid dye SYTO 13 in conjunction with a platelet-specific marker (anti-mouse CD42b-DyLight 649). The efficacy of SYTO 13 for RP identification was confirmed by higher circulating RP levels in platelet-depleted mice during the recovery phase (35% ± 13%) compared to untreated mice (11% ± 1%, n = 9, p < .0001). We further characterized murine RP by exploring the surface expression of the platelet activation marker P-selectin. While there was no preactivation at baseline, stimulation with a thrombin receptor agonist (PAR-4) resulted in a higher increase in the percentage of P-selectin-positive platelets in RP (93% ± 6%) compared to non-RP (77% ± 14%, n = 6, p = .0065). In addition, RP exhibited higher geometric mean fluorescence intensity levels than non-RP (1874 ± 1278 versus 859 ± 549, n = 6, p = .02). Our proof-of-principle study demonstrates the efficacy of SYTO 13 in combination with platelet (activation) markers for identifying RP in both resting and activated states. This method holds promise for simultaneously monitoring RP levels and platelet activation in murine models of human disease.https://www.tandfonline.com/doi/10.1080/09537104.2025.2489020Flow CytometryReticulated PlateletsImmature PlateletsPlatelet SubpopulationsSYTO 13 |
| spellingShingle | Muataz Ali Hamad Krystin Krauel Nancy Schanze Mark Zurek Christoph B. Olivier Peter Kohl Thomas G. Nührenberg Daniel Duerschmied A multi-color flow cytometric method for characterizing murine reticulated platelets using SYTO 13 Platelets Flow Cytometry Reticulated Platelets Immature Platelets Platelet Subpopulations SYTO 13 |
| title | A multi-color flow cytometric method for characterizing murine reticulated platelets using SYTO 13 |
| title_full | A multi-color flow cytometric method for characterizing murine reticulated platelets using SYTO 13 |
| title_fullStr | A multi-color flow cytometric method for characterizing murine reticulated platelets using SYTO 13 |
| title_full_unstemmed | A multi-color flow cytometric method for characterizing murine reticulated platelets using SYTO 13 |
| title_short | A multi-color flow cytometric method for characterizing murine reticulated platelets using SYTO 13 |
| title_sort | multi color flow cytometric method for characterizing murine reticulated platelets using syto 13 |
| topic | Flow Cytometry Reticulated Platelets Immature Platelets Platelet Subpopulations SYTO 13 |
| url | https://www.tandfonline.com/doi/10.1080/09537104.2025.2489020 |
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