Whole-Mount Imaging of Adipose Tissue Macrophages

The adipose tissue comprises highly heterogeneous macrophage populations, which play critical roles in the regulation of adipose tissue function and dysfunction during health and disease. Whole-amount staining is a powerful technique for macrophage characterization within the 3D environment of the adipose tissue, enabling the visualization of different macrophage populations and their interaction with other cells within their in vivo niche. Due to the high-fat content and softness, freezing and sectioning of adipose tissue is difficult, and distortion of tissue morphology typically occurs, especially in the case of white adipose tissue. We describe here a whole-mount staining alternative for adipose tissue imaging that preserves all structures and allows high-resolution image acquisition. We address in a step-by-step manner how to perform immunofluorescence staining of different fat pads and how to optimally visualize cellular and acellular (extracellular matrix) constituents of the adipose tissue and its vasculature, as well as resident and infiltrating macrophage populations.