Laboratório de Imunobiofotônica, Departamento de Morfologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Belo Horizonte, Brasil.
Imunofarmacologia, Departamento de Bioquímica e Imunologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Belo Horizonte, Brasil.
Nat Protoc. 2015 Feb;10(2):258-68. doi: 10.1038/nprot.2015.006. Epub 2015 Jan 8.
Imaging of live animals using intravital microscopy (IVM) has provided a substantial advance in our understanding of cell biology. Here we describe how to adapt a conventional, relatively low-cost laser-scanning microscope to operate as a versatile imaging station. We present the surgical procedures needed to perform liver confocal IVM in mice, thereby allowing one to image different cells in their native environment, including hepatocytes, endothelial cells and leukocytes, as well as to analyze their morphology and function under physiological or pathological conditions. In addition, we propose a plethora of working doses of antibodies and probes to stain multiple cells and molecules simultaneously in vivo. Considering the central role of the liver in metabolism and immunity and the growing interest in the relationship between immune and parenchymal cells, this protocol, in which 20 min of preparation yields up to 4 h of imaging, provides useful insights for various research fields. In addition, the protocol can be easily adapted to investigate adipose tissue, mesentery, intestines, spleen and virtually any abdominal organ.
利用活体显微镜（IVM）对活体动物进行成像，极大地促进了我们对细胞生物学的理解。本文介绍了如何将传统的、相对低成本的激光扫描显微镜改装成多功能成像站。我们还展示了在小鼠肝脏中进行共聚焦 IVM 所需的手术程序，从而可以在其天然环境中对不同细胞进行成像，包括肝细胞、内皮细胞和白细胞，还可以在生理或病理条件下分析它们的形态和功能。此外，我们还提出了大量的抗体和探针工作剂量，可用于在体内同时对多种细胞和分子进行染色。鉴于肝脏在代谢和免疫中的核心作用，以及人们对免疫细胞和实质细胞之间关系的日益关注，本方案仅需 20 分钟的准备，即可获得长达 4 小时的成像时间，为各个研究领域提供了有价值的见解。此外，该方案还可以轻松地用于研究脂肪组织、肠系膜、肠道、脾脏以及几乎任何腹部器官。
