Blocking autofluorescence in brain tissues affected by ischemic stroke, hemorrhagic stroke, or traumatic brain injury
Autofluorescence is frequently observed in animal tissues, interfering with an experimental analysis and leading to inaccurate results. Sudan black B (SBB) is a staining dye widely used in histological studies to eliminate autofluorescence. In this study, our objective was to characterize brain tiss...
| Main Authors: | , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2023-05-01
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| Series: | Frontiers in Immunology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fimmu.2023.1168292/full |
| _version_ | 1797817484864126976 |
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| author | Shaoshuai Wang Shaoshuai Wang Xiuhua Ren Junmin Wang Qinfeng Peng Xiaoyu Niu Chunhua Song Changsheng Li Chao Jiang Weidong Zang Marietta Zille Xiaochong Fan Xuemei Chen Jian Wang Jian Wang |
| author_facet | Shaoshuai Wang Shaoshuai Wang Xiuhua Ren Junmin Wang Qinfeng Peng Xiaoyu Niu Chunhua Song Changsheng Li Chao Jiang Weidong Zang Marietta Zille Xiaochong Fan Xuemei Chen Jian Wang Jian Wang |
| author_sort | Shaoshuai Wang |
| collection | DOAJ |
| description | Autofluorescence is frequently observed in animal tissues, interfering with an experimental analysis and leading to inaccurate results. Sudan black B (SBB) is a staining dye widely used in histological studies to eliminate autofluorescence. In this study, our objective was to characterize brain tissue autofluorescence present in three models of acute brain injury, including collagenase-induced intracerebral hemorrhage (ICH), traumatic brain injury (TBI), and middle cerebral artery occlusion, and to establish a simple method to block autofluorescence effectively. Using fluorescence microscopy, we examined autofluorescence in brain sections affected by ICH and TBI. In addition, we optimized a protocol to block autofluorescence with SBB pretreatment and evaluated the reduction in fluorescence intensity. Compared to untreated, pretreatment with SBB reduced brain tissue autofluorescence in the ICH model by 73.68% (FITC), 76.05% (Tx Red), and 71.88% (DAPI), respectively. In the TBI model, the ratio of pretreatment to untreated decreased by 56.85% (FITC), 44.28% (Tx Red), and 46.36% (DAPI), respectively. Furthermore, we tested the applicability of the protocol using immunofluorescence staining or Cyanine-5.5 labeling in the three models. SBB treatment is highly effective and can be applied to immunofluorescence and fluorescence label imaging techniques. SBB pretreatment effectively reduced background fluorescence but did not significantly reduce the specific fluorescence signal and greatly improved the signal-to-noise ratio of fluorescence imaging. In conclusion, the optimized SBB pretreatment protocol blocks brain section autofluorescence of the three acute brain injury models. |
| first_indexed | 2024-03-13T08:54:08Z |
| format | Article |
| id | doaj.art-35003820fbd346789828262757693dad |
| institution | Directory Open Access Journal |
| issn | 1664-3224 |
| language | English |
| last_indexed | 2024-03-13T08:54:08Z |
| publishDate | 2023-05-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Immunology |
| spelling | doaj.art-35003820fbd346789828262757693dad2023-05-29T04:20:46ZengFrontiers Media S.A.Frontiers in Immunology1664-32242023-05-011410.3389/fimmu.2023.11682921168292Blocking autofluorescence in brain tissues affected by ischemic stroke, hemorrhagic stroke, or traumatic brain injuryShaoshuai Wang0Shaoshuai Wang1Xiuhua Ren2Junmin Wang3Qinfeng Peng4Xiaoyu Niu5Chunhua Song6Changsheng Li7Chao Jiang8Weidong Zang9Marietta Zille10Xiaochong Fan11Xuemei Chen12Jian Wang13Jian Wang14Department of Pain Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, ChinaDepartment of Human Anatomy, School of Basic Medical Sciences, Zhengzhou University, Zhengzhou, Henan, ChinaDepartment of Human Anatomy, School of Basic Medical Sciences, Zhengzhou University, Zhengzhou, Henan, ChinaDepartment of Human Anatomy, School of Basic Medical Sciences, Zhengzhou University, Zhengzhou, Henan, ChinaDepartment of Human Anatomy, School of Basic Medical Sciences, Zhengzhou University, Zhengzhou, Henan, ChinaDepartment of Anesthesiology and Perioperative Medicine, Affiliated Cancer Hospital of Zhengzhou University, Henan, ChinaDepartment of Epidemiology and Statistics, College of Public Health, Zhengzhou University, Zhengzhou, Henan, ChinaDepartment of Anesthesiology and Perioperative Medicine, Affiliated Cancer Hospital of Zhengzhou University, Henan, ChinaDepartment of Neurology, The Fifth Affiliated Hospital of Zhengzhou University, Henan, ChinaDepartment of Human Anatomy, School of Basic Medical Sciences, Zhengzhou University, Zhengzhou, Henan, ChinaDepartment of Pharmaceutical Sciences, Division of Pharmacology and Toxicology, University of Vienna, Vienna, AustriaDepartment of Pain Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, ChinaDepartment of Human Anatomy, School of Basic Medical Sciences, Zhengzhou University, Zhengzhou, Henan, ChinaDepartment of Pain Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, ChinaDepartment of Human Anatomy, School of Basic Medical Sciences, Zhengzhou University, Zhengzhou, Henan, ChinaAutofluorescence is frequently observed in animal tissues, interfering with an experimental analysis and leading to inaccurate results. Sudan black B (SBB) is a staining dye widely used in histological studies to eliminate autofluorescence. In this study, our objective was to characterize brain tissue autofluorescence present in three models of acute brain injury, including collagenase-induced intracerebral hemorrhage (ICH), traumatic brain injury (TBI), and middle cerebral artery occlusion, and to establish a simple method to block autofluorescence effectively. Using fluorescence microscopy, we examined autofluorescence in brain sections affected by ICH and TBI. In addition, we optimized a protocol to block autofluorescence with SBB pretreatment and evaluated the reduction in fluorescence intensity. Compared to untreated, pretreatment with SBB reduced brain tissue autofluorescence in the ICH model by 73.68% (FITC), 76.05% (Tx Red), and 71.88% (DAPI), respectively. In the TBI model, the ratio of pretreatment to untreated decreased by 56.85% (FITC), 44.28% (Tx Red), and 46.36% (DAPI), respectively. Furthermore, we tested the applicability of the protocol using immunofluorescence staining or Cyanine-5.5 labeling in the three models. SBB treatment is highly effective and can be applied to immunofluorescence and fluorescence label imaging techniques. SBB pretreatment effectively reduced background fluorescence but did not significantly reduce the specific fluorescence signal and greatly improved the signal-to-noise ratio of fluorescence imaging. In conclusion, the optimized SBB pretreatment protocol blocks brain section autofluorescence of the three acute brain injury models.https://www.frontiersin.org/articles/10.3389/fimmu.2023.1168292/fullautofluorescenceSudan black Bintracerebral hemorrhagetraumatic brain injuryICHcerebral ischemia |
| spellingShingle | Shaoshuai Wang Shaoshuai Wang Xiuhua Ren Junmin Wang Qinfeng Peng Xiaoyu Niu Chunhua Song Changsheng Li Chao Jiang Weidong Zang Marietta Zille Xiaochong Fan Xuemei Chen Jian Wang Jian Wang Blocking autofluorescence in brain tissues affected by ischemic stroke, hemorrhagic stroke, or traumatic brain injury Frontiers in Immunology autofluorescence Sudan black B intracerebral hemorrhage traumatic brain injury ICH cerebral ischemia |
| title | Blocking autofluorescence in brain tissues affected by ischemic stroke, hemorrhagic stroke, or traumatic brain injury |
| title_full | Blocking autofluorescence in brain tissues affected by ischemic stroke, hemorrhagic stroke, or traumatic brain injury |
| title_fullStr | Blocking autofluorescence in brain tissues affected by ischemic stroke, hemorrhagic stroke, or traumatic brain injury |
| title_full_unstemmed | Blocking autofluorescence in brain tissues affected by ischemic stroke, hemorrhagic stroke, or traumatic brain injury |
| title_short | Blocking autofluorescence in brain tissues affected by ischemic stroke, hemorrhagic stroke, or traumatic brain injury |
| title_sort | blocking autofluorescence in brain tissues affected by ischemic stroke hemorrhagic stroke or traumatic brain injury |
| topic | autofluorescence Sudan black B intracerebral hemorrhage traumatic brain injury ICH cerebral ischemia |
| url | https://www.frontiersin.org/articles/10.3389/fimmu.2023.1168292/full |
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