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Chemical Labeling Hero

Chemical Labeling

Isobaric chemical labeling approaches have enabled a new generation of multi-plexed quantitative proteomic studies. The strategy labels peptides primary amines with isotope-coded mass tags that are indistinguishable from each other during normal mass measurement; however, upon dissociation and tandem mass spectrometry unique reporter ions are produced and used for quantification.  We are developing a new dimethylated leucine (DiLeu) scaffold for isobaric tag technology.  The DiLeu platforms incorporates the same mass defect concepts as our NeuCode labels and has potential to profoundly increase the number of samples that can be co-analyzed in a single LC-MS/MS experiment. 

section under TRD#1 heading

Designed and synthesized in-house, DiLeu isobaric tags are compact and offer excellent performance, often enhancing peptide backbone fragmentation.  DiLeu-tagged peptides therefore can improve confidence in sequence identification all while generating reporter ions at strikingly high abundance. Besides providing a wide quantitative dynamic range, the reporter ion signal affords improved multiplexing capacity, as the overall ion population can be amply divided between extra quantitative channels. And with the ability to synthesize DiLeu cost-effectively, on-demand, and in bulk, experiments of grand scope and scale are well within reach.

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Ultra-plexed Proteome Analysis

 

Ultra-plexed Proteome Analysis

Presently our DiLeu tagging chemistry stands at the forefront of multiplexed labeling approaches by offering parallel analysis of up to twelve proteomes.  To achieve this level of plexing we leverage the slight mass differences in neutron binding energy between carbon, nitrogen, and hydrogen through unique isotopic formulations that differ in mass by as little as 6 mDa (just as in the NeuCode labels of TR&D #2). For twelve-plex DiLeu, twelve discrete reporter ions in the mass range of m/z 115-118 can be distinguished in MS2 scans acquired at a resolving power of 30,000. We are currently developing additional DiLeu isotopologues that differ in mass by 3 mDa to create a 21-plex set of labels and an unparalleled level of quantitative throughput.

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Sample Flexibility and Applications

Sample Flexibility
and Applications

DiLeu isobaric tags feature efficient and convenient amine-reactive chemistry for labeling proteins, peptides, and metabolites derived from a wide variety of biological samples and clinical specimens, ranging from tissue extracts to cerebrospinal fluid or urine. The versatility of DiLeu to label nearly any sample makes it ideal for studies of human disease and mammalian disease models that require compatible proteomic methods to quantify changes within several different types of biological samples, or across multiple experimental conditions, and the reagents can be produced at the scale appropriate for the needs of the study. Currently, research of pancreatic cancer, restenosis, lower urinary tract symptoms, and Alzheimer’s disease are all employing DiLeu and its variants for biological insight.

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DiLeu reagents

Di-Leu Label Reagents

DiLeu reagents serve as attractive alternatives for isobaric tags for relative and absolute quantitation (iTRAQ) and tandem mass tags (TMTs) due to their synthetic simplicity, labeling efficiency, and improved fragmentation efficiency. DiLeu reagent resembles the general structure of a tandem mass tag in that it contains an amine reactive group (triazine ester) targeting the N-terminus and ε-amino group of the lysine side chain of a peptide, a balance group, and a reporter group. For information on this reagent, contact Dr. Lingjun Li through the link below.

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Chemical Labeling Publication Library

Chemical Labeling Publication Library

Ongoing development has led to other viable dimethylated amino acid tags, alternative applications, and novel implementations.
Read more about DiLeu and its variants, including their use for protein, peptide, and metabolite quantification, and much more.

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DiLeuPMP: a multiplexed isobaric labeling method for quantitative analysis of O-glycans

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Recent Advances in Analytical Approaches for Glycan and Glycopeptide Quantitation

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Integrated Label-Free and 10-plex DiLeu Isobaric Tag Quantitative Methods for Profiling Changes in the Mouse Hypothalamic Neuropeptidome and Proteome: Assessment of the Impact of the Gut Microbiome

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Recent Advances in Analytical Approaches for Glycan and Glycopeptide Quantitation

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Mass Defect-based DiLeu Tagging for Multiplexed Data-Independent Acquisition

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Urinary Amine Metabolomics Characterization with Custom 12-plex Isobaric DiLeu Labeling

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Isobaric labeling strategy utilizing 4-plex N,N-dimethyl leucine (DiLeu) tags reveals proteomic changes induced by chemotherapy in cerebrospinal fluid of children with B-cell acute lymphoblastic leukemia

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Urinary metabolomic and proteomic analyses in a mouse model of prostatic inflammation

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21-plex DiLeu Isobaric Tags for High-throughput Quantitative Proteomics

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Spatiotemporal proteomics reveals the molecular consequences of hormone treatment in a mouse model of lower urinary tract dysfunction

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In utero and lactational 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure exacerbates urinary dysfunction in hormone-treated C57BL/6J mice…

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Proteomic modulation in the dorsal spinal cord following spinal cord stimulation therapy in an in vivo neuropathic pain model

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High-Resolution Enabled 5-plex Mass Defect-based N,N-Dimethyl Leucine Tags for Quantitative Proteomics

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In-depth Quantification of Extracellular Matrix Proteins from Human Pancreas

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A Strategy for Discovery and Verification of Candidate Biomarkers in Cerebrospinal Fluid of Preclinical Alzheimer’s Disease

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Metandem: An online software tool for mass spectrometry-basedisobaric labeling metabolomics

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Quantitative Proteomics for Analyses of Multiple Samples in Parallel with Chemical Perturbation

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Quantitative Proteomic Analysis of a Genetically Induced Prostate Inflammation Mouse Model via Custom 4-plex DiLeu Isobaric Labeling

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5-plex Mass Defect-based N,N-Dimethyl Leucine Tags for Proteomics

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In-depth Quantification of Extracellular Matrix Proteins from Human Pancreas

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HOTMAQ Quantification Method for Targeted Proteomics

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Increased N,N-Dimethyl Leucine Isobaric Tag Multiplexing

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Protocol: Proteome analysis with neutron-encoded isotope labeling

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Relative quantitation of neuropeptides using N, N-dimethyl leucine isobaric tags

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Relative quantification of N-glycans

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Site-specific characterization and quantitation of N-glycopeptides (EThcD)

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Expanding DiLeu to Eight Quantitative Channels

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DiLeu Allows for Multiplex Quantification of Urinary Proteins in Men

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Further Multiplexing with Quantitative Proteomics Enabled by Mass Defect and High Resolution MS

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Further Multiplexing with Quantitative Proteomics Enabled by Mass Defect and High Resolution MS

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Absolute Quantification with isotopic DiLeu (iDiLeu)

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Absolute Quantification with isotopic DiLeu (iDiLeu)

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Utilizing DiLeu for Quantifying Amine-Containing Metabolites

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Exploring Other Dimethylated Amino Acids

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A Combined Isobaric and Mass Difference Quantification Strategy (mdDiLeu)

Contact

For additional information about NCQBCS please send your contact details using the form below.
Note, if you are interested in training, please use the form on our training page.

Contact

For additional information about NCQBCS please send your contact details (name, institution, email) along with a short description of how we can help to laura[dot]vantoll[at]wisc[dot]edu.

Note, if you are interested in training, please use the form on our training page.

Coon Lab, University of Wisconsin-Madison, 425 Henry Mall, Madison, WI
Li Lab, University of Wisconsin-Madison, 777 Highland Avenue, Madison, WI
Pagliarini Lab, Washington University School of Medicine, 4523 Clayton Ave, St. Louis, MO