Glycolipids are components of the milk fat globule membrane, and they participate in many biological processes. Among these are the binding of pathogenic microorganisms and their toxins, which leads to the prevention of infection. Glycolipids are difficult to analyze due to their structural complexity. The Glycolipids Project focuses on the development of mass spectrometry-based methods to analyze glycolipids with high throughput and high sensitivity. Matrix-assisted desorption/ionization (MALDI) Fourier transform ion cyclotron resonance (FTICR) mass spectrometry is used to assess the ceramide portion of glycolipids, while tandem mass spectrometry is used to determine the structure of the oligosaccharide or complex sugar portion. A number of common gangliosides in bovine and human milk have been identified including Neu5Ac-Neu5Ac-Hex-Hex-Cer (GD3) and Neu5Ac-Hex-Hex-Cer (GM3). The long-term goal of the Glycolipids Project is to develop the tools necessary to analyze the composition and precise structure of glycolipids and to determine their specific biological functions.