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1.

001-es BibID:BIBFORM084562
035-os BibID:(Scopus)84961282078 (WOS)000379133500002
Első szerző:Bodnár Judit
Cím:Enzymatic removal of N-glycans by PNGase F coated magnetic microparticles / Bodnar Judit, Szekrenyes Akos, Szigeti Marton, Jarvas Gabor, Krenkova Jana, Foret Frantisek, Guttman Andras
Dátum:2016
ISSN:0173-0835 1522-2683
Megjegyzések:Investigation of protein glycosylation is an important area in biomarker discovery and biopharmaceutical research. Alterations in protein N-glycosylation can be an indication of changes in pathological conditions in the medical field or production parameters of biotherapeutics. Rapid development of these disciplines calls for fast, high-throughput, and reproducible methods to analyze protein N-glycosylation. Currently used methods require either long deglycosylation times or large excess of enzymes. In this paper, we report on the use of PNGase F immobilization onto the surface of magnetic microparticles and their use in rapid and efficient removal of N-glycans from glycoproteins. The use of immobilized PNGase F also allowed reusability of the enzyme-coated beads as the magnetic microparticles can be readily partitioned from the sample by a magnet after each deglycosylation reaction. The efficiency and activity of the PNGase F coated magnetic beads was compared with in-solution enzyme reactions using standard glycoproteins possessing the major N-glycan types of neutral, high mannose, and highly sialylated carbohydrates. The PNGase F coated magnetic beads offered comparable deglycosylation level to the conventional in-solution based method in 10-min reaction times for the model glycoproteins of immunoglobulin G (mostly neutral carbohydrates), ribonuclease B (high mannose type sugars), and fetuin (highly sialylated oligosaccharides) with the special features of easy removal of the enzyme from the reaction mixture and reusability.
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
Deglycosylation
Enzyme immobilization
Magnetic microparticles
PNGase F
Megjelenés:Electrophoresis. - 37 : 10 (2016), p. 1264-1269. -
További szerzők:Szekrényes Ákos (1983-) (vegyészmérnök) Szigeti Márton (1986-) (környezetmérnök) Járvás Gábor (1982-) (vegyészmérnök) Krenkova, Jana Foret, František Guttman András (1954-) (vegyészmérnök)
Pályázati támogatás:K116263
NKFIH
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2.

001-es BibID:BIBFORM073661
Első szerző:Borza Beáta (biomérnök)
Cím:Glycosimilarity assessment of biotherapeutics 1 : quantitative comparison of the N-glycosylation of the innovator and a biosimilar version of etanercept / Borza Beata, Szigeti Marton, Szekrenyes Akos, Hajba Laszlo, Guttman Andras
Dátum:2018
ISSN:0731-7085
Megjegyzések:The carbohydrate moieties on the polypeptide chains in most glycoprotein based biotherapeutics and their biosimilars play essential roles in such major mechanisms of actions as antibody-dependent cell-mediated cytotoxicity, complement-dependent cytotoxicity, anti-inflammatory functions and serum clearance. In addition, alteration in glycosylation may influence the safety and efficacy of the product. Glycosylation, therefore, is considered as one of the important critical quality attributes of glycoprotein biotherapeutics, and consequently for their biosimilar counterparts. Thus, the carbohydrate moieties of such biopharmaceuticals (both innovator and biosimilar products) should be closely scrutinized during all stages of the manufacturing process. In this paper we introduce a rapid, capillary gel electrophoresis based process to quantitatively assess the glycosylation aspect of biosimilarity (referred to as glycosimilarity) between the innovator and a biosimilar version of etanercept (Enbrel? and Benepali?, respectively), based on their N-linked carbohydrate profiles. Differences in sialylated, core fucosylated, galactosylated and high mannose glycans were all quantified. Since the mechanism of action of etanercept is TNF? binding, only mannosylation was deemed as critical quality attribute for glycosimilarity assessment due to its influence on serum half-life.
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
Megjelenés:Journal of Pharmaceutical and Biomedical Analysis. - 153 (2018), p. 182-185. -
További szerzők:Szigeti Márton (1986-) (környezetmérnök) Szekrényes Ákos (1983-) (vegyészmérnök) Hajba László Guttman András (1954-) (vegyészmérnök)
Pályázati támogatás:K 116263
NKFIH
GINOP-2.3.2-15-2016-00017
GINOP
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3.

001-es BibID:BIBFORM073868
035-os BibID:(WoS)000428490100015 (Scopus)85040665841
Első szerző:Hajba László
Cím:On the glycosylation aspects of biosimilarity / Hajba László, Szekrényes Ákos, Borza Beáta, Guttman András
Dátum:2018
ISSN:1359-6446
Megjegyzések:The recent expiration of several protein therapeutics opened the door for biosimilar development. Biosimilars are biologic medical products that are similar but not identical copies of already-authorized protein therapeutics. Critical quality attributes (CQA), such as post-translational modifications of recombinant biotherapeutics, are important for the clinical efficacy and safety of both the innovative biologics and their biosimilar counterparts. Here, we summarize biosimilarity CQAs, considering the regulatory guidelines and the statistical aspects (e.g., biosimilarity index) and then discuss glycosylation as one of the important attributes of biosimilarity. Finally, we introduced the 'Glycosimilarity Index', which is based on the averaged biosimilarity criterion.
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
Megjelenés:Drug Discovery Today. - 23 : 3 (2018), p. 616-625. -
További szerzők:Szekrényes Ákos (1983-) (vegyészmérnök) Borza Beáta (1990-) (biomérnök) Guttman András (1954-) (vegyészmérnök)
Pályázati támogatás:K 116263
NKFIH
BIONANO_GINOP-2.3.2-15-2016-00017
GINOP
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4.

001-es BibID:BIBFORM047874
035-os BibID:PMID:21732557
Első szerző:Kovács András László (biológus, biológia-kémia tanár)
Cím:Fractionation of the human plasma proteome for monoclonal antibody proteomics-based biomarker discovery / András Kovács, Edit Sperling, József Lázár, Attila Balogh, János Kádas, Ákos Szekrényes, László Takács, István Kurucz, András Guttman
Dátum:2011
ISSN:0173-0835
Megjegyzések:mAb proteomics, a reversed biomarker discovery approach, is a novel methodology to recognize the proteins of biomarker potential, but requires subsequent antigen identification steps. While in case of high-abundant proteins, it generally does not represent a problem, for medium or lower abundant proteins, the identification step requires a large amount of sample to assure the proper amount of antigen for the ID process. In this article, we report on the use of combined chromatographic and precipitation techniques to generate a large set of fractions representing the human plasma proteome, referred to as the Analyte Library, with the goal to use the relevant library fractions for antigen identification in conjunction with mAb proteomics. Starting from 500mL normal pooled human plasma, this process resulted in 783 fractions with the average protein concentration of 1mg/mL. First, the serum albumin and immunoglobulins were depleted followed by prefractionation by ammonium sulfate precipitation steps. Each precipitate was then separated by size exclusion chromatography, followed by cation and anion exchange chromatography. The 20 most concentrated ion exchange chromatography fractions were further separated by hydrophobic interaction chromatography. All chromatography and precipitation steps were carefully designed aiming to maintain the native forms of the intact proteins throughout the fractionation process. The separation route of vitamin D-binding protein (an antibody proteomics lead) was followed in all major fractionation levels by dot blot assay in order to identify the library fraction it accumulated in and the identity of the antigen was verified by Western blot.
Tárgyszavak:Természettudományok Kémiai tudományok idegen nyelvű folyóiratközlemény külföldi lapban
Megjelenés:Electrophoresis. - 32 : 15 (2011), p. 1916-1925. -
További szerzők:Sperling Edit Lázár József Balogh Attila (bőrgyógyász) Kádas János (1976-) (molekuláris biológus, biokémikus, kertészmérnök) Szekrényes Ákos (1983-) (vegyészmérnök) Takács László (1955-) Kurucz István Guttman András (1954-) (vegyészmérnök)
Pályázati támogatás:K81839
OTKA
TECH-09-A1-2009-0113; mABCHIC
Egyéb
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5.

001-es BibID:BIBFORM050938
Első szerző:Krenkova, Jana
Cím:Oriented immobilization of peptide-N-glycosidase F on a monolithic support for glycosylation analysis / Jana Krenkova, Akos Szekrenyes, Zsolt Keresztessy, Frantisek Foret, Andras Guttman
Dátum:2013
ISSN:0021-9673
Megjegyzések:In this paper, we report on a novel oriented peptide-N-glycosidase F (PNGase F) immobilization approach onto methacrylate based monolithic support for rapid, reproducible and efficient release of the N-linked carbohydrate moieties from glycoproteins. The glutathione-S-transferase-fusion PNGase F (PNGase F-GST) was expressed in Escherichia coli using regular vector technology. The monolithic pore surface was functionalized with glutathione via a succinimidyl-6-(iodoacetyl-amino)-hexanoate linker and the specific affinity of GST toward glutathione was utilized for the oriented coupling. This novel immobilization procedure was compared with reductive amination technique commonly used for non-oriented enzyme immobilization via primary amine functionalities. Both coupling approaches were compared using enzymatic treatment of several glycoproteins, such as ribonuclease B, fetuin and immunoglobulin G followed by MALDI/MS and CE-LIF analysis of the released glycans. Orientedly immobilized PNGase F via GST-glutathione coupling showed significantly higher activity, remained stable for several months, and allowed rapid release of various types of glycans (high-mannose, core fucosylated, sialylated, etc.) from glycoproteins. Complete protein deglycosylation was obtained as fast as in several seconds when using flow-through immobilized microreactors.
Tárgyszavak:Természettudományok Kémiai tudományok idegen nyelvű folyóiratközlemény külföldi lapban
Enzyme microreactor
Oriented immobilization
Monolith
PNGase F
Deglycosylation
Megjelenés:Journal of Chromatography A. - 1322 (2013), p. 54-61. -
További szerzők:Szekrényes Ákos (1983-) (vegyészmérnök) Keresztessy Zsolt Foret, František Guttman András (1954-) (vegyészmérnök)
Pályázati támogatás:K-81839
OTKA
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6.

001-es BibID:BIBFORM050953
Első szerző:Smejkal, Petr
Cím:Chip-based CE for rapid separation of 8-aminopyrene-1,3,6-trisulfonic acid (APTS) derivatized glycans / Petr Smejkal, Ákos Szekrényes, Markéta Ryvolová, František Foret, András Guttman, Fritz Bek, Mirek Macka
Dátum:2010
ISSN:0173-0835
Megjegyzések:Fluorescently labeled carbohydrates released from glycoproteins were separated using a commercially available microfluidic chip electrophoresis system. While the instrumentation was primarily designed for DNA analysis it was found that the application base can be easily expanded using the development software provided by the manufacturer. The carbohydrates were released by enzymatic digestion (PNGase F) from glycoproteins present in human plasma after boronic acid ? lectin affinity enrichment. After fluorescent labeling with 8-aminopyrene-1,3,6-trisulfonic acid the carbohydrates were separated based on capillary gel electrophoresis mechanism and detected by a fluorescence detector using a blue (470?nm) LED. The separation was completed in 40?s in a microfluidic channel of 14?mm length. Glucose ladder carbohydrate oligomers differing by one glucose unit were baseline separated up to a 20-mer with the main limitation being the detection sensitivity. As expected, the observed resolution in these experiments did not approach that of standard CE with 20 times longer separation distance; however, the chip-based analysis excelled in the speed of the separation. Similar electrophoretic profiles of glycans released from plasma glycoproteins were obtained using a standard CE equipment with 35?cm separation length and microfluidic chips with a separation distance of only 14?mm.
Tárgyszavak:Természettudományok Kémiai tudományok idegen nyelvű folyóiratközlemény külföldi lapban
APTS
Bioanalyzer
Chip-based analysis
Glycans
PSA
Megjelenés:Electrophoresis. - 31 : 22 (2010), p. 3783-3786. -
További szerzők:Szekrényes Ákos (1983-) (vegyészmérnök) Ryvolová, Markéta Foret, František Guttman András (1954-) (vegyészmérnök) Bek, Fritz Macka, Mirek
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7.

001-es BibID:BIBFORM065041
Első szerző:Székely Andrea
Cím:Multi Capillary SDS-Gel Electrophoresis for the Analysis of Fluorescently Labeled mAb preparations: a high throughput quality control process for the production of QuantiPlasma and PlasmaScan mAb libraries / Andrea Székely, Ákos Szekrényes, Márta Kerékgyártó, Attila Balogh, János Kádas, József Lázár, András Guttman, István Kurucz, László Takács
Dátum:2014
ISSN:0173-0835
Megjegyzések:Molecular heterogeneity of mAb preparations is the result of various co- and post-translational modifications and to contaminants related to the production process. Changes in molecular composition results in alterations of functional performance, therefore quality control and validation of therapeutic or diagnostic protein products is essential. A special case is the consistent production of mAb libraries (QuantiPlasma? and PlasmaScan?) for proteome profiling, quality control of which represents a challenge because of high number of mAbs (>1000). Here, we devise a generally applicable multicapillary SDS-gel electrophoresis process for the analysis of fluorescently labeled mAb preparations for the high throughput quality control of mAbs of the QuantiPlasma? and PlasmaScan? libraries.
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
capillary electrophoresis
glycoform
mAb proteomics
quality control
SDS
Megjelenés:Electrophoresis. - 35 : 15 (2014), p. 2155-2162. -
További szerzők:Szekrényes Ákos (1983-) (vegyészmérnök) Kerékgyártó Márta (1987-) (molekuláris biológus) Balogh Attila Kádas János (1976-) (molekuláris biológus, biokémikus, kertészmérnök) Lázár József Guttman András (1954-) (vegyészmérnök) Kurucz István Takács László (1955-)
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8.

001-es BibID:BIBFORM084931
Első szerző:Szekrényes Ákos (vegyészmérnök)
Cím:Multi-Site N-glycan mapping study 1: Capillary electrophoresis ? laser induced fluorescence / Szekrényes Ákos, Park SungAe Suhr, Santos Marcia, Lew Clarence, Jones Aled, Haxo Ted, Kimzey Michael, Pourkaveh Shiva, Szabó Zoltán, Sosic Zoran, Feng Peng, Váradi Csaba, de l'Escaille François, Falmagne Jean-Bernard, Sejwal Preeti, Niedringhaus Thomas, Michels David, Freckleton Gordon, Hamm Melissa, Manuilov Anastasiya, Schwartz Melissa, Luo Jiann-Kae, van Dyck Jonathan, Leung Pui-King, Olajos Marcell, Gu Yingmei, Gao Kai, Wang Wenbo, Wegstein Jo, Tep Samnang, Guttman András
Dátum:2016
ISSN:1942-0862 1942-0870
Megjegyzések:An international team that included 20 independent laboratories from biopharmaceutical companies, universities, analytical contract laboratories and national authorities in the United States, Europe and Asia was formed to evaluate the reproducibility of sample preparation and analysis of N-glycans using capillary electrophoresis of 8-aminopyrene1,3,6-trisulfonic acid (APTS)-labeled glycans with laser induced fluorescence (CE-LIF) detection (16 sites) and ultra highperformance liquid chromatography (UHPLC, 12 sites; results to be reported in a subsequent publication). All participants used the same lot of chemicals, samples, reagents, and columns/capillaries to run their assays. Migration time, peak area and peak area percent values were determined for all peaks with >0.1% peak area. Our results demonstrated low variability and high reproducibility, both, within any given site as well across all sites, which indicates that a standard N-glycan analysis platform appropriate for general use (clone selection, process development, lot release, etc.) within the industry can be established.
Tárgyszavak:Orvostudományok Klinikai orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
Biotherapeutics
Capillary electrophoresis
Intercompany study
N-glycans
Megjelenés:mAbs. - 8 : 1 (2016), p. 56-64. -
További szerzők:Park, SungAe Suhr Santos, Marcia Lew, Clarence Jones, Aled Haxo, Ted Kimzey, Michael Pourkaveh, Shiva Szabó Zoltán Sosic, Zoran Feng, Peng Váradi Csaba (1988-) (molekuláris biológus) de l'Escaille, François Falmagne, Jean-Bernard Sejwal, Preeti Niedringhaus, Thomas Michels, David Freckleton, Gordon Hamm, Melissa Manuilov, Anastasiya Schwartz, Melissa Luo, Jiann-Kae van Dyck, Jonathan Leung, Pui-King Olajos Marcell Gu, Yingmei Gao, Kai Wang, Wenbo Wegstein, Jo Tep, Samnang Guttman András (1954-) (vegyészmérnök)
Pályázati támogatás:K 116263
NKFIH
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9.

001-es BibID:BIBFORM083126
035-os BibID:(WoS)000508387300020 (Scopus)85075579456
Első szerző:Szekrényes Ákos (vegyészmérnök)
Cím:Quantitative comparison of the N-glycosylation of therapeutic glycoproteins using the Glycosimilarity Index. A tutorial / Szekrenyes Akos, Szigeti Marton, Dvorakova Veronika, Jarvas Gabor, Guttman Andras
Dátum:2020
ISSN:0165-9936
Megjegyzések:Glycosylation is considered as one of the crucial critical quality attributes of therapeutic glycoproteins and for their biosimilar counterparts. Carbohydrate moieties of such biopharmaceuticals should be closely monitored during all stages of development and manufacturing and studied accordingly during comparability or similarity exercises. In therapeutic glycoprotein production, the batch-to-batch alteration in glycosylation represents an excellent indicator of process robustness. Due to the complexity of the obtained glycoanalytical profile it is challenging to determine and describe the level of similarity between the N-glycosylation characteristics of two glycoprotein production lots or between the reference batch and its biosimilar versions. In this tutorial, we provide a step-by-step approach to calculate a numerical similarity value of N-glycosylation, referred to as Glycosimilarity Index that can be efficiently used to assess the conformity degree of any new N-glycosylation profile to a given target profile.
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
Megjelenés:Trac-Trends In Analytical Chemistry. - 122 (2020), p. 1-8. -
További szerzők:Szigeti Márton (1986-) (környezetmérnök) Dvorakova, Veronika Járvás Gábor (1982-) (vegyészmérnök) Guttman András (1954-) (vegyészmérnök)
Pályázati támogatás:BIONANO_GINOP-2.3.2-15-2016-00017
GINOP
K116263
NKFIH
NN127062
NKFIH
2018-2.1.17-TET-KR-2018-00010
NKFIH
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10.

001-es BibID:BIBFORM077852
035-os BibID:(PMID)29330871 (Scopus)85041568961 (WOS)000429417500010
Első szerző:Szekrényes Ákos (vegyészmérnök)
Cím:Multi-site N-Glycan mapping study 2 : UHPLC / Ákos Szekrényes, SungAe Suhr Park, Eoin Cosgrave, Aled Jones, Ted Haxo, Michael Kimzey, Shiva Pourkaveh, Zoltán Szabó, Zoran Sosic, Peng Feng, Preeti Sejwal, Kelsey Dent, David Michels, Gordon Freckleton, Jun Qian, Catherine Lancaster, Toni Duffy, Melissa Schwartz, Jiann-Kae Luo, Jonathan van Dyck, Pui-King Leung, Marcell Olajos, Ronald Kowle, Kai Gao, Wenbo Wang, Jo Wegstein, Samnang Tep, Apolka Domokos, Csaba Váradi, András Guttman
Dátum:2018
ISSN:0173-0835 1522-2683
Megjegyzések:In the first part of this publication, the results from an international study evaluating theprecision (i.e., repeatability and reproducibility) of N-glycosylation analysis using capillaryelectrophoresis of APTS-labeled N-glycans were presented. The corresponding resultsfrom ultra-high performance liquid chromatography (UHPLC) with fluorescence detectionare presented here from 12 participating sites. All participants used the same lot of samples,reagents, and columns to perform the assays. Elution time, peak area and peak areapercent values were determined for all peaks 0.1% peak area, and statistical analysis wasperformed following ISO 5725-2 guideline principles. The results demonstrated adequatereproducibility, within any given site as well across all sites, indicating that standardUHPLC-based N-glycan analysis platforms are appropriate for general use
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
2-AB
Biotherapeutics
Intercompany study
N-glycans
UHPLC
Megjelenés:Electrophoresis. - 39 : 7 (2018), p. 998-1005. -
További szerzők:Park, SungAe Suhr Cosgrave, Eoin Jones, Aled Haxo, Ted Kimzey, Michael Pourkaveh, Shiva Szabó Zoltán (orvos) Sosic, Zoran Feng, Peng Sejwal, Preeti Dent, Kelsey Michels, David Freckleton, Gordon Qian, Jun Lancaster, Catherine Duffy, Toni Schwartz, Melissa Luo, Jiann-Kae van Dyck, Jonathan Leung, Pui-King Olajos Marcell Kowle, Ronald Gao, Kai Wang, Wenbo Wegstein, Jo Tep, Samnang Domokos Apolka Váradi Csaba (1988-) (molekuláris biológus) Guttman András (1954-) (vegyészmérnök)
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11.

001-es BibID:BIBFORM058348
Első szerző:Szekrényes Ákos (vegyészmérnök)
Cím:Sample preparation for N-glycosylation analysis of therapeutic monoclonal antibodies by electrophoresis / Ákos Szekrényes, Jan Partyka, Csaba Varadi, Jana Krenkova, Frantisek Foret, András Guttman
Dátum:2015
Megjegyzések:There are a considerable number of biopharmaceuticals that have been approved for clinical use in the past decade. Over half of these new generation drugs are glycoproteins, such as monoclonal antibodies or other recombinant glycoproteins, which are mostly produced in mammalian cell lines. The linked carbohydrate moieties affect not only their physicochemical properties and thermal stability but also crucial features like receptor-binding activity, circulating half-life, as well as immunogenicity. The structural diversity of these attached glycans can be manifested in altered monosaccharide composition and linkages/positions among the monosaccharide building blocks. In addition, as more and more biosimilar products hit the market, understanding the effects of their glycosylation modification has become a recent target in efficacy and safety issues. To ensure consistent quality of these products, glycosylation profiles have to be monitored and controlled in all steps of the manufacturing process, i.e., from clone selection to lot release. In this paper, we describe some of the recently introduced and commonly used sample preparation techniques for capillary electrophoresis (CE)-based profiling and structural elucidation of N-glycans. The presented protocols include protein A affinity partitioning of monoclonal antibodies (mAbs), enzymatic release of the N-linked glycans, labeling of the liberated carbohydrates, reaction mixture purification techniques to remove the excess labeling reagent, and high-resolution and rapid capillary electrophoresis-laser-induced fluorescence (CE-LIF)-based profiling of the labeled and purified N-glycans.
ISBN:978-1-4939-2353-3
Tárgyszavak:Orvostudományok Elméleti orvostudományok könyvfejezet
N-glycosylation analysis
Megjelenés:Microchip Capillary Electrophoresis Protocols : Methods in Molecular Biology / Ann Van Schepdael. - p. 183-195. -
További szerzők:Partyka, Jan Váradi Csaba (1988-) (molekuláris biológus) Krenkova, Jana Foret, František Guttman András (1954-) (vegyészmérnök)
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12.

001-es BibID:BIBFORM050948
Első szerző:Szekrényes Ákos (vegyészmérnök)
Cím:High-throughput analysis of therapeutic and diagnostic monoclonal antibodies by multicapillary SDS gel electrophoresis in conjunction with covalent fluorescent labeling / Ákos Szekrényes, Udo Roth, Márta Kerékgyártó, Andrea Székely, István Kurucz, Karen Kowalewski, András Guttman
Dátum:2012
ISSN:1618-2642
Megjegyzések:Capillary gel electrophoresis (CGE) in the presence of sodium dodecyl sulfate (SDS) is a well-established and widely used protein analysis technique in the biotechnology industry, and increasingly becoming the method of choice that meets the requirements of the standards of International Conference of Harmonization (ICH). Automated single channel capillary electrophoresis systems are usually equipped with UV absorbance and/or laser-induced fluorescent (LIF) detection options offering general applicability and high detection sensitivity, respectively; however, with limited throughput. This shortcoming is addressed by the use of multicapillary gel electrophoresis (mCGE) systems with LED-induced fluorescent detection (LED-IF), also featuring automation and excellent detection sensitivity, thus widely applicable to rapid and large-scale analysis of biotherapeutics, especially monoclonal antibodies (mAb). The methodology we report in this paper is readily applicable for rapid purity assessment and subunit characterization of IgG molecules including detection of non-glycosylated heavy chains (NGHC) and separation of possible subunit variations such as truncated light chains (Pre-LC) or alternative splice variants. Covalent fluorophore derivatization and the mCGE analysis of the labeled IgG samples with multi-capillary gel electrophoresis are thoroughly described. Reducing and non-reducing conditions were both applied with and without peptide N-glycosidase F mediated deglycosylation.
Tárgyszavak:Természettudományok Kémiai tudományok idegen nyelvű folyóiratközlemény külföldi lapban
Biotherapeutics
Quality analysis
Multicapillary gel electrophoresis
Antibody purity
Fluorescent labeling
Megjelenés:Analytical and Bioanalytical Chemistry. - 404 : 5 (2012), p. 1485-1494. -
További szerzők:Roth, Udo Kerékgyártó Márta (1987-) (molekuláris biológus) Székely Andrea Kurucz István Kowalewski, Karen Guttman András (1954-) (vegyészmérnök)
Pályázati támogatás:K-81839
OTKA
TECH-09-A1-2009-0113
Egyéb
Internet cím:Szerző által megadott URL
DOI
Intézményi repozitóriumban (DEA) tárolt változat
Borító:
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