Analysis of photoreceptor rod outer segment phagocytosis by RPE cells in situ.
Analysis of photoreceptor rod outer segment phagocytosis by RPE cells in situ.
复制标题
DOI：
10.1007/978-1-62703-080-9_17
复制
发表时间：
2013-01-01
期刊：
影响因子：
--
通讯作者：
Finnemann, Silvia C
中科院分区：
文献类型：
--
作者：
Sethna, Saumil;Finnemann, Silvia C
在一天中的不同时间，通过对视网膜色素上皮(RPE)中视紫红质阳性吞噬小体的计数，可以定量评估白天RPE吞噬作用的吞噬和消化阶段，从而有效地清除神经视网膜中脱落的光感受器外片段(POS)。比较年龄和背景相匹配的实验野生型和突变小鼠或大鼠的这种活动有助于确定特定蛋白质在吞噬过程中的作用。在这里，我们描述了小鼠眼睛的采集、包埋、切片、RPE细胞中杆状POS噬菌体的免疫荧光标记、用激光扫描共聚焦显微镜对RPE中的POS噬菌体进行成像以及POS定量的实验过程。
Counting rhodopsin-positive phagosomes residing in the retinal pigment epithelium (RPE) in the eye at different times of day allows a quantitative assessment of engulfment and digestion phases of diurnal RPE phagocytosis, which efficiently clears shed photoreceptor outer segment fragments (POS) from the neural retina. Comparing such activities among age- and background-matched experimental wild-type and mutant mice or rats serves to identify roles for specific proteins in the phagocytic process. Here, we describe experimental procedures for mouse eye harvest, embedding, sectioning, immunofluorescence labeling of rod POS phagosomes in RPE cells in sagittal eye sections, imaging of POS phagosomes in the RPE by laser scanning confocal microscopy, and POS quantification.