Lipids are essential for numerous cellular functions and signaling processes. However, they are understudied compared to other biological molecules such as proteins mainly due to a lack of appropriate methodology. Compared to proteins, it is very difficult to introduce perturbations such as concentration changes and it is further very challenging to visualize lipids especially in a cellular context. This has mainly to do with both the size and also the chemical diversity of lipids. Chemical biology approaches such as the introduction of photolabile moieties, photocrosslinking- and click chemistry-groups offer the possibility to fill this methodological gap and in particular to address biological questions on the level of individual lipid species. Using diacylglycerols as an exemplary lipid class, we employed such an approach to understand how individual lipid species can exert a specific function in the cell and how minor structural differences can influence lipid dynamics in the plasma membrane.