Comparison of procainamide and 2-aminobenzamide labeling for profiling and identification of glycans by liquid chromatography with fluorescence detection coupled to electrospray ionization–mass spectrometry

Kozak, Radoslaw P.; Tortosa, Concepcion Badia; Fernandes, Daryl L.; Spencer, Daniel I. R.

doi:10.1016/j.ab.2015.06.006

Cited by 106 publications

(74 citation statements)

References 12 publications

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“…Of the released O-glycans, 40% was fluorescently labelled with procainamide as described previously [43] using Ludger procainamide glycan labelling kit containing 2-picoline borane. Briefly, samples in 10 μL of water were mixed with 10 μL of procainamide labelling solution and incubated at 65°C for 1 h. The samples were evaporated to dryness by vacuum centrifugation and re-suspended in water (100 μL) for further analysis.…”

Section: Methodsmentioning

confidence: 99%

Variation of Human Salivary O-Glycome

et al. 2016

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The study of saliva O-glycosylation is receiving increasing attention due to the potential of glycans for disease biomarkers, but also due to easy access and non-invasive collection of saliva as biological fluid. Saliva is rich in glycoproteins which are secreted from the bloodstream or produced by salivary glands. Mucins, which are highly O-glycosylated proteins, are particularly abundant in human saliva. Their glycosylation is associated with blood group and secretor status, and represents a reservoir of potential disease biomarkers. This study aims to analyse and compare O-glycans released from whole human mouth saliva collected 3 times a day from a healthy individual over a 5 days period. O-linked glycans were released by hydrazinolysis, labelled with procainamide and analysed by ultra-high performance liquid chromatography with fluorescence detection (UHPLC-FLR) coupled to electrospray ionization mass spectrometry (ESI-MS/MS). The sample preparation method showed excellent reproducibility and can therefore be used for biomarker discovery. Our data demonstrates that the O-glycosylation in human saliva changes significantly during the day. These changes may be related to changes in the salivary concentrations of specific proteins.

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Section: Methodsmentioning

confidence: 99%

Variation of Human Salivary O-Glycome

et al. 2016

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“…Tertiary amine groups are usually utilized in the tags, for example, procainamide (ProA), 14, 15 N 2 , N 2 , N 4 , N 4 -tetraethyl-6-hydrazinyl-1, 3, 5-triazine-2, 4-diamine (Meladrazine) 16 and RapiFluor-MS (RFMS) 17 all contain tertiary amines for increased ionization efficiency in positive ion mode. Quantitative glycomics can also be achieved using multiplexing reagent derivatization.…”

Section: Introductionmentioning

confidence: 99%

Direct comparison of derivatization strategies for LC-MS/MS analysis ofN-glycans

Zhou

Veillon

Dong

et al. 2017

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Protein glycosylation is a common post-translational modification that has significant impacts on protein folding, lifespan, conformation, distribution and function. N-glycans, which are attached to asparagine residues of proteins, are studied most often due to their compatibility with enzymatic release. Despite the ease of N-glycan release, compositional and structural complexity coupled with poor ionization efficiency during liquid chromatography mass spectrometry (LC-MS) make quantitative glycomic studies a significant challenge. To overcome these challenges, glycans are almost always derivatized prior to LC-MS analyses to impart favorable characteristics, such as improved ionization efficiency, increased LC separation efficiency and the production of more informative fragments during tandem MS. There are a number of derivatization methods available for LC-MS analysis of glycans, each of which imparts different properties that affect both glycan retention on LC columns and MS analyses. To provide guidance for the proper selection of derivatizing reagents and LC columns, herein, we describe a comprehensive assessment of 2-aminobenzamide, procainamide, aminoxyTMT, RapiFluor-MS (RFMS) labeling, reduction and reduction with permethylation for N-glycan analysis. Of the derivatization strategies examined, RFMS provided the highest MS signal enhancement for neutral glycans, while permethylation significantly enhanced the MS intensity and structural stability of sialylated glycans.

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“…Released N‐glycans were fluorescently labeled by reductive amination with procainamide as described previously using the LudgerTagTM Procainamide Glycan Labelling Kit (LT‐KPROC‐24). Procainamide labeled samples were analyzed by HILIC‐(U)HPLC‐ESI‐MS with fluorescence detection.…”

Section: Methodsmentioning

confidence: 99%

Disruption of sialic acid metabolism drives tumor growth by augmenting CD8⁺ T cell apoptosis

Cornelissen

Blanas

Horst

et al. 2019

Intl Journal of Cancer

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Sialylated glycan structures are known for their immunomodulatory capacities and their contribution to tumor immune evasion. However, the role of aberrant sialylation in colorectal cancer and the consequences of complete tumor desialylation on anti‐tumor immunity remain unstudied. Here, we report that CRISPR/Cas9‐mediated knock out of the CMAS gene, encoding a key enzyme in the sialylation pathway, in the mouse colorectal cancer MC38 cell line completely abrogated cell surface expression of sialic acids (MC38‐Sianull) and, unexpectedly, significantly increased in vivo tumor growth compared to the control MC38‐MOCK cells. This enhanced tumor growth of MC38‐Sianull cells could be attributed to decreased CD8+ T cell frequencies in the tumor microenvironment only, as immune cell frequencies in tumor‐draining lymph nodes remained unaffected. In addition, MC38‐Sianull cells were able to induce CD8+ T cell apoptosis in an antigen‐independent manner. Moreover, low CMAS gene expression correlated with reduced recurrence‐free survival in a human colorectal cancer cohort, supporting the clinical relevance of our work. Together, these results demonstrate for the first time a detrimental effect of complete tumor desialylation on colorectal cancer tumor growth, which greatly impacts the design of novel cancer therapeutics aimed at altering the tumor glycosylation profile.

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Comparison of procainamide and 2-aminobenzamide labeling for profiling and identification of glycans by liquid chromatography with fluorescence detection coupled to electrospray ionization–mass spectrometry

Cited by 106 publications

References 12 publications

Variation of Human Salivary O-Glycome

Variation of Human Salivary O-Glycome

Direct comparison of derivatization strategies for LC-MS/MS analysis ofN-glycans

Disruption of sialic acid metabolism drives tumor growth by augmenting CD8⁺ T cell apoptosis

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Comparison of procainamide and 2-aminobenzamide labeling for profiling and identification of glycans by liquid chromatography with fluorescence detection coupled to electrospray ionization–mass spectrometry

Cited by 106 publications

References 12 publications

Variation of Human Salivary O-Glycome

Variation of Human Salivary O-Glycome

Direct comparison of derivatization strategies for LC-MS/MS analysis ofN-glycans

Disruption of sialic acid metabolism drives tumor growth by augmenting CD8+ T cell apoptosis

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Disruption of sialic acid metabolism drives tumor growth by augmenting CD8⁺ T cell apoptosis