High-performance anion-exchange chromatography of oligosaccharides using pellicular resins and pulsed amperometric detection

Townsend, R. Reid; Hardy, Mark R.; Hindsgaul, Ole; Lee, Yuan Chuan

doi:10.1016/0003-2697(88)90044-9

Cited by 164 publications

(60 citation statements)

References 17 publications

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“…6), and their basic structure was found to be galactose(Gal)2 mannose(Man)3. N-acetyl glucosamine(GlcNAc)4 as reported by Townsend et al (25) (see the legend to Fig. 6).…”

Section: Resultsmentioning

confidence: 57%

“…4 nmol each of the aberrant Aa (139-148) peptide and normal fibrinogen was treated with 1 Ml of glycopeptidase A (20 Mg/Mi) as described (22), and the reaction mixtures were directly applied on a CarboPac PA-l column of the Bio-LC system by high performance anion-exchange column chromatography with pulsed-amperometric detection (Dionex Co., Sunnyvale, CA) essentially according to Townsend et al (25).…”

Section: Methodsmentioning

confidence: 99%

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Fibrinogen Lima: a homozygous dysfibrinogen with an A alpha-arginine-141 to serine substitution associated with extra N-glycosylation at A alpha-asparagine-139. Impaired fibrin gel formation but normal fibrin-facilitated plasminogen activation catalyzed by tissue-type plasminogen activator.

Maekawa

Yamazumi²,

Muramatsu³

et al. 1992

J. Clin. Invest.

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An Aa-arginine-141 to serine substitution has been identified in a homozygous dysfibrinogen, fibrinogen Lima, associated with impaired fibrin polymerization. The point mutation created an asparagine-X-serine-type glycosylation sequence, and indeed, extra, mainly disialylated biantennary oligosaccharides have been isolated from Aa asparagine-139 of the patient's fibrinogen. This type of glycosylation sequence is unique for human fibrinogen, because the sequences shown for normal and abnormal fibrinogens are all asparagine-X-threonine types. The terminal sialic acids of the extra oligosaccharides seem to have largely contributed to the impaired fibrin gel formation, as evidenced by its correction to a near normal level by desialylation. Nevertheless, the polymerizing fibrin facilitated tissuetype plasminogen activator-catalyzed plasmin formation in a normal fashion, indicating that the initial two-stranded fibrin protofibrils had been constructed normally. Thus the impaired fibrin gel formation could be attributed to the delay in their subsequent lateral association, most probably because of the repulsive forces generated by the negative electric charge of the extra sialic acids. The substitution of a basic residue arginine to a noncharged residue serine may also have contributed to the impaired function in a similar manner or by steric hindrance in association with bulky extra oligosaccharide chains. (J. Clin. Invest. 1992. 90:67-76.) Key words: amino acid substitutioncongenital homozygous dysfibrinogen * extra asparagine-linked oligosaccharide * fibrin-facilitated plasminogen activation * lateral association of fibrin protofibrils

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“…6), and their basic structure was found to be galactose(Gal)2 mannose(Man)3. N-acetyl glucosamine(GlcNAc)4 as reported by Townsend et al (25) (see the legend to Fig. 6).…”

Section: Resultsmentioning

confidence: 57%

Section: Methodsmentioning

confidence: 99%

Fibrinogen Lima: a homozygous dysfibrinogen with an A alpha-arginine-141 to serine substitution associated with extra N-glycosylation at A alpha-asparagine-139. Impaired fibrin gel formation but normal fibrin-facilitated plasminogen activation catalyzed by tissue-type plasminogen activator.

Maekawa

Yamazumi²,

Muramatsu³

et al. 1992

J. Clin. Invest.

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“…Increasing glycan size and sialylation is believed to cause reduction in PAD response. 68,69 This might account for the small differences for G0F and the sialylated species.…”

Section: Discussionmentioning

confidence: 99%

Comparison of methods for the analysis of therapeutic immunoglobulin G Fc-glycosylation profiles—Part 1: Separation-based methods

Reusch

Haberger

Maier

et al. 2015

mAbs

142

119

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(2015) Comparison of methods for the analysis of therapeutic immunoglobulin G Fc-glycosylation profiles-Part 1: Separation-based methods, mAbs, 7:1, 167-179, DOI: 10.4161/19420862.2014.986000 To link to this article: https://doi.org/10. 4161/19420862.2014 Abbreviations: mAb, monoclonal antibody; Fc, fragment crystallizable; IgG, immunoglobulin G; HILIC-UPLC, hydrophilic interaction liquid chromatography-ultra high performance liquid chromatography; 2-AB, 2-aminobenzamide; Fab, fragment, antigen-binding; CE-LIF, capillary electrophoresis-laser induced fluorescence; HPLC, high performance liquid chromatography; MALDI-MS, matrix-assisted laser desorption/ionization-mass spectrometry; ESI-MS, electrospray ionization-mass spectrometry; HPAEC-PAD, high-performance anion exchange chromatography with pulsed amperometric detection; APTS, 8-aminopyrene-1, 3, 6-trisulfonic acid; DSA-FACE, DNA-sequencer-aided fluorophore-assisted carbohydrate electrophoresis; ANTS, 8-aminonaphthalene-1, 3, 6-trisulfonate; CCGE, cartridge-based capillary gel electrophoresis; HR, high resolution; IAB, InstantAB labeling; CHO, Chinese hamster ovaryImmunoglobulin G (IgG) crystallizable fragment (Fc) glycosylation is crucial for antibody effector functions, such as antibody-dependent cell-mediated cytotoxicity, and for their pharmacokinetic and pharmacodynamics behavior. To monitor the Fc-glycosylation in bioprocess development, as well as product characterization and release analytics, reliable techniques for glycosylation analysis are needed. A wide range of analytical methods has found its way into these applications. In this study, a comprehensive comparison was performed of separation-based methods for Fcglycosylation profiling of an IgG biopharmaceutical. A therapeutic antibody reference material was analyzed 6-fold on 2 different days, and the methods were compared for precision, accuracy, throughput and other features; special emphasis was placed on the detection of sialic acid-containing glycans. Seven, non-mass spectrometric methods were compared; the methods utilized liquid chromatography-based separation of fluorescent-labeled glycans, capillary electrophoresis-based separation of fluorescent-labeled glycans, or high-performance anion exchange chromatography with pulsed amperometric detection. Hydrophilic interaction liquid chromatography-ultra high performance liquid chromatography of 2-aminobenzamide (2-AB)-labeled glycans was used as a reference method. All of the methods showed excellent precision and accuracy; some differences were observed, particularly with regard to the detection and quantitation of minor glycan species, such as sialylated glycans.

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“…Anion exchange with pulsed amperometric detection (HPAEC-PAD) has been used for analysis of acidic oligosaccharides by three groups. Townsend et al (1988) separated the oligosaccharides on the basis of the number of sialic acids per molecule at pH 4.6, and further resolved them by desialylation and resolution of the resulting neutral oligosaccharides at pH 13 [23]. This two-step process overcame the inconsistent response factors for sialyloligosaccharides with different linkages by the electrochemical detector.…”

Section: Discussionmentioning

confidence: 99%

“…Native acidic oligosaccharides of milk have been resolved into monosialyl-, disialyl-, and trisialyloligosaccharide mixtures by mediumpressure anion-exchange chromatography with absorption at 214 nm [22], but further resolution was not achieved. A technique more commonly employed for native milk oligosaccharide resolution and quantification is high pH anion exchange chromatography with pulsed amperometric detection [17,[23][24][25][26], but the electrochemical response factors for anionic sialyloligosaccharides by this technique are not consistent, and differ by the linkage position of the sialic acid [23]. Reverse-or normal-phase chromatography was able to resolve derivatized or reduced acidic oligosaccharides, with detection by UV absorbance or spectrofluorimetry as the basis for quantification [27,28].…”

Section: Introductory Statementmentioning

confidence: 99%

Simultaneous quantification of sialyloligosaccharides from human milk by capillary electrophoresis

Bao

Zhu

Newburg

2007

Analytical Biochemistry

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The acidic oligosaccharides of human milk are predominantly sialyloligosaccharides. Pathogens that bind sialic acid-containing glycans on their host mucosal surfaces may be inhibited by human milk sialyloligosaccharides, but testing this hypothesis requires their reliable quantification in milk. Sialyloligosaccharides have been quantified by anion exchange HPLC, reverse or normal phase HPLC, and capillary electrophoresis (CE) of fluorescent derivatives; in milk, these oligosaccharides have been analyzed by high pH anion exchange chromatography with pulsed amperometric detection, and, in our laboratory, by CE with detection at 205 nm. The novel method described herein uses a running buffer of aqueous 200 mM NaH 2 PO 4 at pH 7.05 containing 100 mM SDS made 45% (v/v) with methanol to baseline resolve five oligosaccharides, and separate all 12. This allows automated simultaneous quantification of the 12 major sialyloligosaccharides of human milk in a single 35-minute run. This method revealed differences in sialyloligosaccharide concentrations between less and more mature milk from the same donors. Individual donors also varied in expression of sialyloligosaccharides in their milk. Thus, the facile quantification of sialyloligosaccharides by this method is suitable for measuring variation in expression of specific sialyloligosaccharides in milk and their relationship to decreased risk of specific diseases in infants. Keywordshuman milk oligosaccharides; sialyloligosaccharides; capillary electrophoresis INTRODUCTORY STATEMENTHuman milk contains a large number of compounds that provide nutritional support and contribute toward defense of the newborn. Human milk oligosaccharides (HMOS), a principal component of human milk, are found in concentrations of 6 to 12 g/L, and these oligosaccharides vary in size, structure, and abundance [1]. The oligosaccharides of milks from other species vary widely, and the composition of oligosaccharides from human milk is unique [2].HMOS play a potentially protective role to nursing infants. Breastfed infants have a different microbiota than infants fed artificially, and human milk glycans are thought to act as prebiotics that promote the growth of Bifidobacterium bifidus [3]. Breast-fed infants have fewer or less severe gastrointestinal and respiratory infections during the first year of life than formula-fed

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High-performance anion-exchange chromatography of oligosaccharides using pellicular resins and pulsed amperometric detection

Cited by 164 publications

References 17 publications

Fibrinogen Lima: a homozygous dysfibrinogen with an A alpha-arginine-141 to serine substitution associated with extra N-glycosylation at A alpha-asparagine-139. Impaired fibrin gel formation but normal fibrin-facilitated plasminogen activation catalyzed by tissue-type plasminogen activator.

Fibrinogen Lima: a homozygous dysfibrinogen with an A alpha-arginine-141 to serine substitution associated with extra N-glycosylation at A alpha-asparagine-139. Impaired fibrin gel formation but normal fibrin-facilitated plasminogen activation catalyzed by tissue-type plasminogen activator.

Comparison of methods for the analysis of therapeutic immunoglobulin G Fc-glycosylation profiles—Part 1: Separation-based methods

Simultaneous quantification of sialyloligosaccharides from human milk by capillary electrophoresis

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