The enucleated eyes were fixated for 10 minutes in neutral formalin buffer 10% (CellPath Ltd., Newton Powys, UK). A frontal cut was made at the ora serrata, and the anterior part of the eye was removed. The remaining posterior eye cup was fixated in neutral formalin buffer at room temperature overnight. The area containing the laser application was identified under a light microscope (Leica CLS, 10X; Leica, Wetzlar, Germany), and an area extending at least 3 mm on each side of the lesion was separated to a depth including both the retina and the choroid using a surgical knife (disposable scalpel; Swann-Morton, Sheffield, UK). After dehydration and paraffin embedding, 4 µm sections were cut, and every tenth section was stained with hematoxylin and eosin and mounted on glass slides. The sections containing the laser application were identified in light microscope (VS120 Virtual Slide Microscope; Olympus, Tokyo, Japan) and were documented with its inbuilt camera (CellSense Dimension; Olympus). For each section containing a part of the laser application, the following section was selected for immunohistochemistry. After deparaffination in Xylen (Merck, Darmstadt, Germany), each of these sections were placed in citrate-buffer (2.1 g citrate monohydrate/l, pH 6,0) and were heated in a microwave oven at 85°C for 3 × 3 minutes. The sections were placed on a rocking table for 30 minutes in order to adapt to room temperature, and were washed 3 times with phosphate buffered saline (PBS; pH 7,4) and incubated with PBS + 2% BSA for 10 minutes. Subsequently, the sections were incubated with the endothelial cell marker Griffonia Simplicifolia Lectin I, isolectin B4 (GSL I, isolectin B4; Vector Laboratories, Burlingame, CA, USA; diluted 1:50 in PBS +1% BSA) at 4°C overnight. The following day, the sections were washed 3 times in PBS at room temperature, stained with 4′,6-diamidino-2-phenylindole (DAPI; 2 mg/mL; Sigma, St. Louis, MO, USA), rinsed twice in distilled water, dipped in ethanol, and dried. LifterSlip premium printed cover glasses (Erie Scientific Company, Portsmouth, NH, USA) were applied using ProLong Gold antifade reagent (Life Technologies Corporation, Eugene, OR, USA). The sections were examined by light microscope and lesions documented by fluorescence microscopy (VS120 Virtual Slide Microscope; Olympus) using the cellSense software for image capture and processing.