Quantitative Proteomics of Xenopus Embryos I, Sample Preparation. Author Meera Gupta, Matthew Sonnett, Lillia Ryazanova, Marc Presler, Martin Wühr Publication Year 2018 Type Journal Article Abstract Xenopus oocytes and embryos are model systems optimally suited for quantitative proteomics. This is due to the availability of large amount of protein material and the ease of physical manipulation. Furthermore, facile in vitro fertilization provides superbly synchronized embryos for cell cycle and developmental stages. Here, we detail protocols developed over the last few years for sample preparation of multiplexed proteomics with TMT-tags followed by quantitative mass spectrometry analysis using the MultiNotch MS3 approach. In this approach, each condition is barcoded with an isobaric tag at the peptide level. After barcoding, samples are combined and the relative abundance of ~100,000 peptides is quantified on a mass spectrometer. High reproducibility of the sample preparation process prior to peptides being tagged and combined is of upmost importance for obtaining unbiased data. Otherwise, differences in sample handling can inadvertently appear as biological changes. We detail and exemplify the application of our sample workflow on an embryonic time-series of ten developmental stages of Xenopus laevis embryos ranging from the egg to stage 35 (just before hatching). Our accompanying paper (Chapter 14 ) details a bioinformatics pipeline to analyze the quality of the given sample preparation and strategies to convert spectra of X. laevis peptides into biologically interpretable data. Keywords Animals, Mass Spectrometry, Hydrogen-Ion Concentration, Chromatography, Liquid, Proteomics, Xenopus Proteins, Embryo, Nonmammalian, Cysteine, Peptides, Xenopus, Quality Control, Chemical Precipitation, Egg Yolk, Solid Phase Extraction Journal Methods Mol Biol Volume 1865 Pages 175-194 Date Published 2018 ISSN Number 1940-6029 DOI 10.1007/978-1-4939-8784-9_13 Alternate Journal Methods Mol Biol PMCID PMC6564683 PMID 30151767 PubMedPubMed CentralGoogle ScholarBibTeXEndNote X3 XML