## 2 Investigating In Vivo Stroke Model Using Photochemical Thrombosis  
### Photochemical Thrombosis-Based Stroke Animal Model Establishments  
Each of research, diagnosis and treatment modalities require verification, especially preclinical and clinical tests, which are mandatory for diagnosis and treatment devices that are to be employed for medical purposes. Development of a stroke-induced animal model for preclinical tests and modalities to build such an animal model are being extensively investigated [16; 17; 18; 19]. A typical stroke animal model investigation is achieved either through surgical blocking of specific vessels [20; 21; 22] or through a biochemical method [23; 24]. However, stroke in local brain regions is investigated using photochemical thrombosis. An investigation of a photochemical stroke animal model is based on a generation of blood slots using rose Bengal [25; 26; 27; 28]. Rose Bengal reacts with a green light irradiation to produce reactive oxygen species, which then activate factors that are involved in a coagulation of blood and production of thrombosis. To use this characteristic of rose Bengal, an animal's blood vessel is first injected with rose Bengal, and after waiting for it to spread to all of the blood vessels, a green light is irradiated in a desired brain region, which causes photochemical thrombosis and stroke in the desired area. This section describes several studies focused on photochemical thrombosis investigation modalities for performance enhancements such as an improvement of location accuracy of brain damages and an inhibition of excessive brain damage. [PERSON] et al. investigated a photothrombosis inducing module that was integrated with an upright microscope to generate a photothrombotic stroke on a single brain arteriole [29]. To be specific, a 532-nm laser diode module and an optical cage system were applied for a construction of the photothrombosis inducing module, which was connected to a filter slot of an upright microscope. Moreover, this module provides a function that can form blood clots in certain cerebral blood vessels while observing them under a microscope. When laser irradiation on a single brain vessel in mice was maintained for 120 s, a simultaneous blockage of a targeted brain vessel was verified using microscopic images (Figure 1a). In addition, laser speckle imaging was utilized to confirm the vascular obstruction caused by photothrombotic stroke. For a similar purpose, [PERSON] et al. developed a reconfigured laser system to reduce a deviation in induced sizes of brain damages caused by photothrombosis [30]. In the laser system that they proposed, a polarizer or a variable neutral density filter were employed to maintain a constant output irradiation power of laser, which was verified using an optical power meter. An in vivo study on photothrombosis inducing in rats' brains and 2,3,5-Triphenyltetrazolzolium chloride (TTC) staining used for estimating locations and depths of photothrombotic strokes confirmed that the system with a maintained optical irradiation power is useful for inducing a stroke and the consequent brain damage of a certain size. [PERSON] et al. investigated a minor photothrombotic stroke animal model using a fiber-optic laser irradiation probe to generate Rose Bengal photoactivation [31]. In detail, a probe with a halogen lamp consisting of an optical fiber with a 13-mm diameter, a heat filter, and collimator was constructed to induce a photothrombotic stroke. According to magnetic resonance images described in Figure 1b, an investigation of a small animal model with minor stroke was achieved using a fiber-optic photothrombosis inducing probe. A photothrombosis inducing device integrated with a stereotaxic frame was also developed to produce brain damage at a certain position [32; 33]. The stereotaxic frame, which was applied in this study, was magnetic resonance compatible (i.e., it was configured to move with an animal's location to acquire magnetic resonance images of the brain).