PDF(Pubmed)
Abstract：
Energy storage and endocrine functions of the Drosophila fat body make it an excellent model for elucidating mechanisms that underlie physiological and pathophysiological organismal metabolism. Combined with Drosophila\'s robust genetic and immunofluorescence microscopy toolkits, studies of Drosophila fat body function are ripe for cell biological analysis. Unlike the larval fat body, which is easily removed as a single, cohesive sheet of tissue, isolating intact adult fat body proves to be more challenging, thus hindering consistent immunofluorescence labeling even within a single piece of adipose tissue. Here, we describe an improved approach to handling Drosophila abdomens that ensures full access of the adult fat body to solutions generally used in immunofluorescence labeling protocols. In addition, we assess the quality of fluorescence reporter expression and antibody immunoreactivity in response to variations in fixative type, fixation incubation time, and detergent used for cellular permeabilization. Overall, we provide several recommendations for steps in a whole-mount staining protocol that results in consistent and robust immunofluorescence labeling of the adult Drosophila fat body.
摘要：
果蝇脂肪体的能量储存和内分泌功能使其成为阐明生理和病理生理有机体代谢基础的机制的极好模型。结合果蝇强大的遗传和免疫荧光显微镜工具包，果蝇脂肪体功能的研究已经成熟，可以进行细胞生物学分析。与幼体脂肪不同，它很容易作为一个单一的移除，粘性组织片,隔离完整的成人脂肪身体被证明更具挑战性，从而阻碍了一致的免疫荧光标记，即使在一片脂肪组织。这里,我们描述了一种处理果蝇腹部的改进方法，该方法可确保成年脂肪体完全接触免疫荧光标记方案中常用的溶液。此外,我们评估了荧光报告表达的质量和抗体免疫反应性，以响应固定剂类型的变化，固定孵育时间，和用于细胞渗透的洗涤剂。总的来说,我们为整个mount染色方案中的步骤提供了一些建议，该方案可对成年果蝇脂肪体进行一致且可靠的免疫荧光标记。
