China
Africa
Explore chapters and articles related to this topic
The Precision Medicine Approach in Oncology
Published in David E. Thurston, Ilona Pysz, Chemistry and Pharmacology of Anticancer Drugs, 2021
MALDI-MSI (Matrix-Assisted Laser Desorption Ionization MS Imaging) and LCM-MS (Laser Capture Microdissection MS) are two new MS-based technologies that are being developed for use in the proteomics area. These techniques, which are still experimental, attempt to use mass spectrometry to image proteins and peptides in cells and tissues. Gel-free isotopic labeling methods such as SILAC (Stable Isotope Labeling with Amino Acids in Cell Culture), iTRAQ (Isobaric Tags for Relative and Absolute Quantitation) and I-TMTs (Isobaric Tandem Mass Tags) are currently capable of quantifying up to thousands of proteins in a single analysis with high reproducibility.
Radiobiology and Hadron Therapy
Published in Manjit Dosanjh, Jacques Bernier, Advances in Particle Therapy, 2018
Eleanor A. Blakely, Manjit Dosanjh
It is not surprising that the novel clustered characteristics of particle-induced DNA damage triggers a unique set of DDR post-lesion formation-signaling cascades and cell cycle arrest and recruitment of DNA repair factors compared to X-ray damage. A systematic study to decipher acute signaling events induced by different radiation qualities using high-resolution mass spectrometry-based proteomics has recently been published by Winter et al. (2017). Two hours after exposure of stable isotope labeling by amino acids in cell culture (SILAC)-labeled human lung adenocarcinoma A549 cells to X-rays, protons or carbon ion showed extensive alterations of the phosphorylation status despite protein expression remaining largely unchanged. Phosphorylation events were similar for proton and carbon irradiation; however, a distinctly different number of sites responded differentially for X-rays. The results were also validated with targeted spike-in experiments. This information will provide unique insight into the differential regulation of phosphorylation sites for radiations of different quality and how they may be further optimised for cancer radiotherapy.
Metabolomics and Proteomics
Published in Crystal D. Karakochuk, Kyly C. Whitfield, Tim J. Green, Klaus Kraemer, The Biology of the First 1,000 Days, 2017
Richard D. Semba, Marta Gonzalez-Freire
Currently, it is possible to measure >10,000 proteins in a single biological sample, something that was not feasible several years ago. These advances are due not only to new innovations in mass spectrometry instrumentation, but also to improvements in sample preparation and standardized guidelines for sample collection. The depletion of highly abundant proteins, such as albumin and immunoglobulins, facilitates the detection of low abundant proteins (the “deep proteome”). More effective electrophoresis and chromatography protocols and tools, such as stable isotope labeling with amino acids in cell culture (SILAC), isobaric tags for relative and absolute quantitation (iTRAQ), and tandem mass tags (TMTs), have facilitated the quantification of proteins [13–15]. Methods are also improving for the detection of posttranslation modifications (PTMs), such as phosphorylation, glycosylation, acetylation, ubiquitination, sumoylation, and citrullination. PTMs are important to study since they reflect the diversity of protein function. Many PTMs are difficult to study because they are labile to sample processing and mass spectrometry. For example, O-GlcNAcylation, an important PTM that rivals phosphorylation in abundance and distribution, has been especially challenging to detect and measure. Many proteins have functions that are unknown or not well understood. By studying the proteins with which a particular protein interacts, it is possible to deduce biological functions and pathways. Protocols have recently been developed for proteomic analysis of dried blood spots and formalin-fixed, paraffin-embedded tissues [16].
Understanding the effect of carrier proteomes in single cell proteomic studies - key lessons
Published in Expert Review of Proteomics, 2022
Pankaj Dwivedi, Christopher M. Rose
Before the introduction of isobaric labels, stable isotope labeling by amino acids in cell culture (SILAC) was the most common multiplexed quantitative proteomic technique [16]. However, SILAC requires a number of cell doubling events to ensure heavy amino acids are sufficiently incorporated to the cellular proteome and enable quantification. Due to this, SILAC analysis of human tissue was not feasible and SILAC analysis of model systems (e.g. mouse models) was possible – but costly. This led researchers to develop methods to analyze tissue samples that circumvented this challenge. Here, a SILAC mix, termed super-SILAC, was developed as a spike-in internal standard [17] (Figure 2). A super-SILAC mix is a mixture of two or more labeled cell lines labeled with heavy amino acids that closely represent the experimental proteome being analyzed (Figure 2). The super-SILAC approach is potentially the first experimental approach to introduce a carrier proteome strategy specifically aimed at enhancing the peptide identification and quantitation of low-level peptides and proteins. More recently, isobaric labels have become the preferred reagents to enable multiplexed quantification due to the increased level of multiplexing compared to SILAC (i.e. 18-plex with isobaric labels vs. 3-plex with SILAC), and the concept of a carrier proteome has evolved within these new multiplexing approaches.
The development and clinical applications of proteomics: an Indian perspective
Published in Expert Review of Proteomics, 2020
Khushman Taunk, Bhargab Kalita, Vaikhari Kale, Venkatesh Chanukuppa, Tufan Naiya, Surekha M. Zingde, Srikanth Rapole
Stable isotope labeling by amino acids in cell culture (SILAC), is a metabolic labeling strategy that was designed to quantify differential protein expression in normal and diseased cells in a cell culture setup. The differentially labeled proteins are isolated, digested with trypsin, and the peptides analyzed by mass spectrometry. The intensities of the labeled and unlabeled peptides provide a measure of their relative abundance [47]. Apart from metabolic labeling, chemical labeling approaches including Isobaric tags for relative and absolute quantification (iTRAQ) and Tandem mass tags (TMT) are also widely used in clinical proteomics studies in which proteolytic peptides are labeled with different isobaric tags [48,49]. Thereafter, labeled peptides are pooled and either directly subjected to mass-spectrometry analysis or fractionated using strong cation exchange chromatography before protein identification. These chemical labeling methods rely on relative intensities of the reporter ions generated via fragmentation. Several clinical proteomics investigations carried out in laboratories from different corners of the country employed different labeling approaches to understand diverse disease pathogenesis and for discovery proteomics studies [50–54].
Global views of proteasome-mediated degradation by mass spectrometry
Published in Expert Review of Proteomics, 2019
It has been known for several decades that there are proteins that are degraded by the proteasome at a rate dependent on the age of the protein, where recently translated proteins were shown to have a faster turnover rate than long-lived ones [10]. However, whether these are underlying principles and whether they hold true for different classes of proteins remained unclear as these studies were done in bulk on specific proteins and could not provide the necessary resolution for proteome-wide analysis. Improvements in MS analysis have provided tremendous capabilities in measurements of protein half-life by improving the depth and sensitivity of analyzing complex biological samples. For example, quantitative analyses that employ metabolic labeling of proteins using stable isotope labeling by amino acids in cell culture (SILAC) allowed for better quantification of protein abundances than MS without labeling [11]. A combination of SILAC with pulse-labeling following proteasome inhibition identified the specific rates of degradation for thousands of cellular proteins [12]. One observation emerging from this study is that many of the proteins with age-dependent degradation were found to be subunits of cellular protein complexes, positing that the incorporation of a protein into a complex may stabilize it. Thus, the ubiquitin-proteasome system is involved in balancing the cellular environment and removing non-stoichiometric subunits from the cell.