Ultra-high sensitivity mass spectrometry quantifies single-cell proteome changes upon perturbation.

Single-cell technologies are revolutionizing biology but are today mainly limited to imaging and deep sequencing. However, proteins are the main drivers of cellular function and in-depth characterization of individual cells by mass spectrometry (MS)-based proteomics would thus be highly valuable and complementary. Here, we develop a robust workflow combining miniaturized sample preparation, very low flow-rate chromatography, and a novel trapped ion mobility mass spectrometer, resulting in a more than 10-fold improved sensitivity. We precisely and robustly quantify proteomes and their changes in single, FACS-isolated cells. Arresting cells at defined stages of the cell cycle by drug treatment retrieves expected key regulators. Furthermore, it highlights potential novel ones and allows cell phase prediction. Comparing the variability in more than 430 single-cell proteomes to transcriptome data revealed a stable-core proteome despite perturbation, while the transcriptome appears stochastic. Our technology can readily be applied to ultra-high sensitivity analyses of tissue material, posttranslational modifications, and small molecule studies from small cell counts to gain unprecedented insights into cellular heterogeneity in health and disease

Calorimetric and Microstructural Investigation of Frozen Hydrated Gluten

The thermal and microstructural properties of frozen hydrated gluten were studied by using differential scanning calorimetry (DSC), modulated DSC, and low-temperature scanning electron microscopy (cryo-SEM). This work was undertaken to investigate the thermal transitions observed in frozen hydrated gluten and relate them to its microstructure. The minor peak that is observed just before the major endotherm (melting of bulk ice) was assigned to the melting of ice that is confined to capillaries formed by gluten. The Defay–Prigogine theory for the depression of melting point of fluids confined in capillaries was put forward in order to explain the calorimetric results. The pore radius size of the capillaries was calculated by using four different empirical models. Kinetic analysis of the growth of the pore radius size revealed that it follows first-order kinetics. Cryo-SEM observations revealed that gluten forms a continuous homogeneous and not fibrous network. Results of the present investigation showed that is impossible to assign a T g value for hydrated frozen gluten because of the wide temperature range over which the gluten matrix vitrifies, and therefore the construction of state diagrams is not feasible at subzero temperatures for this material. Furthermore, the gluten matrix is deteriorated with two different mechanisms from ice recrystallization, one that results from the growth of ice that is confined in capillaries and the other from the growth of bulk ice