Preparation and Negative Staining for Visualization of Cyanoglobule Lipid Droplets Using Transmission Electron Microscopy.

Lipid droplets have emerged as dynamic organelles involved in diverse cellular processes beyond simple lipid storage. In plants and cyanobacteria, growing evidence highlights their importance in stress adaptation and signaling, yet methods to study their structure and purity remain limited. Traditionally, in situ transmission electron microscopy (TEM) has been used to visualize lipid droplets within intact cells. While powerful, this approach cannot easily evaluate isolated lipid droplets or confirm their purity. In this protocol, we describe a rapid method for preparing and visualizing cyanoglobule lipid droplets isolated from cyanobacteria. The isolated droplets are directly processed for TEM using negative staining with uranyl acetate, providing a straightforward and efficient workflow. The procedure can be applied broadly to lipid droplets from diverse organisms, independent of species or cellular origin. This protocol offers a simple, fast, and widely applicable approach to assessing lipid droplets, expanding the toolkit for researchers studying their structure and function. Key features • Provides a brief and detailed method to visualize cyanoglobule lipid droplets. • Offers a rapid workflow (~1-2 h from preparation to imaging), enabling efficient sample processing and high-throughput analysis. • Employs negative staining and TEM to directly assess droplet morphology and purity without complex sample preparation. • Applicable to isolated lipid droplets from diverse organisms, making it a broadly useful tool beyond cyanobacteria.