Thin‐Layer Chromatography and Coomassie Staining of Phospholipids for Fast and Simple Lipidomics Sample Preparation

Abstract Lipids play major roles in basic cellular functions. Their analysis, therefore, gained importance; however, it is complicated by the high complexity of natural lipidomes. To overcome this challenge, pre‐separation of the lipids by thin‐layer chromatography (TLC) and high‐resolution mass spectrometry (MS) are often employed. Here, we explore Coomassie staining for TLC‐separated phospholipids and provide an extraction protocol of stained lipids for subsequent MS analysis. We exemplify our approach by analyzing lipid mixtures varying in complexity and found that TLC pre‐separation increases the number of identified lipid species and lipid classes. In addition, we identify and quantify lipids from polymer nanodiscs. In summary, Coomassie staining of TLC‐separated lipids is well‐suited for phospholipids, is compatible with MS, does not require specialized equipment and can be performed independently of subsequent MS experiments. We envision many future applications of our workflow.