For several years, the spotlight has been on omega-3 very-long-chain polyunsaturated fatty acids (n-3 VLC-PUFAs), such as eicosapentaenoic acid (20:5Δ5,8,11,14,17 n-3, EPA) and docosahexaenoic acid (22:6Δ4,7,10,13,16,19 n-3, DHA) because of their properties beneficial to human health. However, mammalian lipid metabolism is unable to produce n-3 VLC-PUFAs due to the lack of key enzymes allowing their synthesis. A nutritional intake of fish oils or seafood rich in n-3 VLC-PUFAs is a possible alternative, but fish oils can be prone to various contaminations and overfishing is also a major problem. Furthermore, these marine organisms are enriched in n-3 VLC-PUFAs due to their own consumption of phytoplankton, which, at the base of the microbial food web, have the enzymatic machinery necessary for the synthesis of n-3 VLC-PUFAs. These n-3 VLC-PUFAs produced by microalgae accumulate mainly in membrane glycerolipids and betaine lipids and more rarely in oils in the form of lipid droplets (LDs) rich in triacylglycerols (TAGs). These LDs accumulate in microalgae when growing conditions become unfavorable (nutrient deficiency, etc.), impacting biomass and photosynthetic activity. Nowadays, the main effort by biotech companies is to succeed in obtaining microalgae strains capable of accumulating n-3 VLC-PUFAs within TAGs without affecting vital physiological parameters. This chapter summarizes the current knowledge on the metabolism of n-3 VLC-PUFAs in microalgae and provides an overview of the importance of these high-value lipids for food and feed.