脑切片 TTC 染色与梗死面积测定
Browse experimental protocolsFresh brains are cut to 2-mm coronal sections, stained per well with 2% TTC in 0.1M PBS at 37°C for 30 min protected from light, stored in fixative, imaged on a stereomicroscope, and infarct percentage computed ipsilateral-versus-contralateral.
Scope and method
Sample：新鲜小鼠或大鼠脑（取脑后可直接用于 TTC 检测；本方案不含缺血造模步骤）
- middle cerebral artery occlusion spontaneously hypertensive rat
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- permanent middle cerebral artery occlusion with transient common carotid artery occlusion neonatal Wistar rat
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- bilateral common carotid artery occlusion combined with middle cerebral artery occlusion rat
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- direct mechanical middle cerebral artery occlusion rat (strain not reported)
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- middle cerebral artery occlusion rodent (surgical technique category)
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- embolic middle cerebral artery occlusion mouse (strain not reported)
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- distal middle cerebral artery occlusion mouse (strain not reported)
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- permanent middle cerebral artery occlusion mouse (strain not reported)
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- endothelin-1 induced focal cerebral ischemia rodent (species and strain not reported)
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- thromboembolic stroke rodent (species and strain not reported)
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- photothrombotic stroke rodent (species and strain not reported)
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- intraluminal filament middle cerebral artery occlusion rat (strain not reported)
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- intraluminal filament middle cerebral artery occlusion mouse (strain not reported)
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- microsphere embolic stroke rat
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- middle cerebral artery occlusion Swiss mouse
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- permanent distal middle cerebral artery occlusion C57BL/6J mouse
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- photothrombotic cortical stroke mouse
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- embolic middle cerebral artery occlusion rat
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- photothrombotic stroke rat
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- transient middle cerebral artery occlusion Wistar rat
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- thromboembolic stroke rat
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- permanent middle cerebral artery occlusion C57BL/6 mouse
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- endothelin-1 induced focal cerebral ischemia (rat)
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- transient middle cerebral artery occlusion C57BL/6J mouse
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- C57 小鼠 MCAO 线栓模型
- Age
- 2 月龄；体重 20–22 g
- Sex
- Male
- Tissue
- 脑组织
- SD 大鼠 MCAO 线栓模型
- Age
- 成年（来源未报告具体周龄）
- Tissue
- 脑组织（2 mm 冠状切片；梗死区与存活区）
- Koizumi 入路大脑中动脉阻塞模型（啮齿类）
- Tissue
- brain sections
- Zea Longa 入路大脑中动脉阻塞模型（啮齿类）
- Tissue
- brain sections
- 线栓性短暂大脑中动脉阻塞模型（啮齿类）
- Tissue
- brain sections
- 小鼠大脑中动脉局部凝血酶血栓模型
- Tissue
- brain sections
Reagents and materials
Procedure
TTC Staining
TTC Staining The brain is removed from the skull and placed into a brain slicer mold placed in ice. Razor blades are positioned in the mold to obtain 2mm thick sections. Sections are placed in individual wells of a 24 well tissue culture dish with 500 μl of 2% TTC in 0.1M PBS. The plate is wrapped in foil and placed on a 37°C slide warmer for 30 minutes. TTC is removed and 500ul of fixative is added to each well for storage. Images of the sections are captured by Olympus S2H10 research stereo- microscope and Adobe Photoshop software. The infarct area and total area of the contralateral and ipsilateral hemispheres are measured with Scion image software (Scion Corporation). The percent infarct area is calculated as a percentage of the ipsilateral hemisphere relative to the contralateral hemisphere as previously described. [F2.JPG]]()
Quality control and readouts
切片厚度 2 mm；每孔 500 μl 2% TTC（0.1M PBS）；锡箔遮光、37°C 摊片机孵育 30 分钟；吸除 TTC 后每孔 500ul 固定液保存；面积测量按患侧/对侧半球分别勾画后计算百分比。
存活脑组织染为红色、梗死区呈白色；以 2 mm 连续冠状切片逐孔染色成像后，测量患侧与对侧半球的梗死面积及总面积，计算患侧相对对侧的梗死面积百分比。
References
- Ian Simpson. Induce Stroke via Hypoxia-Ischemia. protocols.io, v2, 2019. DOI: 10.17504/protocols.io.8e5htg6
- Hu Y, Yang Z-H, Yan F, et al. CCA repair or ECA ligation - Which middle cerebral artery occlusion is better in the reperfusion mouse model? Ibrain. 2023;9(3):258-269. DOI:10.1002/ibra.12128; PMID:37786756.
- Bederson JB, Pitts LH, Germano SM, Nishimura MC, Davis RL, Bartkowski HM. Evaluation of 2,3,5-triphenyltetrazolium chloride as a stain for detection and quantification of experimental cerebral infarction in rats. Stroke. 1986;17(6):1304-1308. DOI:10.1161/01.STR.17.6.1304; PMID:2433817. 原文以成年大鼠 MCA 闭塞后 TTC 与 H&E 对比，24 小时 TTC 梗死面积 21±2% 的冠状切片面积，r=0.91，支持 TTC 用于大鼠脑梗死检测与定量。
- Clark WM, Lessov N, Lauten JD, Hazel K. Doxycycline treatment reduces ischemic brain damage in transient middle cerebral artery occlusion in the rat. J Mol Neurosci. 1997;9(2):103-108. DOI:10.1007/BF02736854; PMID:9407391. 原文在成年 Sprague-Dawley 大鼠以线栓行 2 小时可逆 MCAO，48 小时以梗死体积为终点（对照 158±44 mm3），支持 SD 大鼠 MCAO 梗死体积检测背景。