To determine the proteins in the membrane raft, we used the TMT-labeling and nano-liquid chromatography mass spectrometry (nano-LC-MS/MS) analysis by Creative Proteomics (NY, USA; https://www.creative-proteomics.com/). Rat primary microglia were treated with IL-6 (25 ng/ml) for 15 min. Membrane rafts were obtained by flotation assay. Samples were prepared from three independent experiments. Proteins in equal volumes of raft fractions were digested with trypsin, desalted, and labeled with a TMT reagent (Thermo Fisher Science). The TMT-labeled peptides were fractionated and analyzed by nano-LC-MS/MS. The resulting MS/MS data were analyzed and searched against the rat protein database using Proteome Discoverer 2.1.

The exploitation of computational techniques to predict the outcome of chemical reactions is becoming commonplace, enabling a reduction in the number of physical experiments required to optimize a reaction. Here, we adapt and combine models for polymerization kinetics and molar mass dispersity as a function of conversion for reversible addition fragmentation chain transfer (RAFT) solution polymerization, including the introduction of a novel expression accounting for termination. A flow reactor operating under isothermal conditions was used to experimentally validate the models for the RAFT polymerization of dimethyl acrylamide with an additional term to accommodate the effect of residence time distribution. Further validation is conducted in a batch reactor, where a previously recorded in situ temperature monitoring provides the ability to model the system under more representative batch conditions, accounting for slow heat transfer and the observed exotherm. The model also shows agreement with several literature examples of the RAFT polymerization of acrylamide and acrylate monomers in batch reactors. In principle, the model not only provides a tool for polymer chemists to estimate ideal conditions for a polymerization, but it can also automatically define the initial parameter space for exploration by computationally controlled reactor platforms provided a reliable estimation of rate constants is available. The model is compiled into an easily accessible application to enable simulation of RAFT polymerization of several monomers.

Selected Hits from Mass Spectrometry Assays from Lipid Rafts Proteins.