Articles by Maria Ekemohn

Introducing our new CFO, Johny Humaloja

September 9, 2019 | Genovis Team, Interview |

 

Johny joined the Genovis team in August, and he brings more than 25 years of financial experience to the company. We are very excited to have you in the team, Johny! Welcome!

 

Tell us a bit about yourself

I live in Malmö, in the southern part of Sweden and I have two daughters. I have a cat named Zelda. On my free time I like to play hockey with my friends and go out for a ride on my road bike.

 

If you were to describe yourself using only one word – what word would that be?

Committed

 

Tell us about your previous working life

I have more than 18 years of finance leadership experience in the life science industry, mainly from working in US public listed companies such as Biogen and Boston Scientific. Focus has been on the Nordic region, to develop businesses and execute operational improvements. At Biogen I got the opportunity to work as Plant Controller and run all the financial activities at their large-scale manufacturing plant in North Carolina.

 

What will your main focus be here at Genovis?

Day-to-day business and make sure to enable Genovis to continue to grow and expand the business model. It will be key for me to understand the different parts of the organization and give adequate financial information and support. Financial areas to focus on will be;internal control, project management, statutory accounting, taxes, manage outsourced services, and external reporting.

 

What do you believe will be the biggest opportunity in your new position as CFO?

I believe I can utilize skills and experiences in sales/commercial and production area to find new ways to do business and provide financial and strategic leadership.

 

4 quick questions:

Coffee or tea?

Coffee – Zoegas

Aerosmith or Depeche Mode?

Depeche Mode

Ice cream or candy?

Chocolate Ice cream

Soccer or ice hockey?

Hockey, Carolina Hurricanes

FragIT™ kit in Study Evaluating IgG Charge

September 2, 2019 | References |

 

Protein charge is a fundamental property that influences both the ability to interact with other molecules and the structure, solubility and stability of the protein.

In this collaborative work by Boehringer Ingelheim Pharmaceuticals, Janssen BioTherapeutics and University of New Hampshire, the charge measurements of twelve monoclonal IgG and their F(ab’)2 and Fc fragments are presented. Besides other interesting findings, it is suggested that mAb charge measurements is valuable when it comes to selecting candidate molecules for development.

FragIT kit was used to achieve the IgG subunits before charge evaluation. This kit consists of spin columns of an immobilized version of the FabRICATOR (IdeS) enzyme for antibody digestion and spin columns for affinity binding of the Fc fragments. Using this kit, the Fc and F(ab’)2 fragments were easily separated from each other, with no enzyme in the final preparation.

 

For more information about FragIT kit, please visit the following page:

FragIT kit

 

The full-text paper is available online:

Yang et al., 2019. IgG Charge: Practical and Biological Implications. Antibodies, 8(1), 24

 

The Digested Guide to ASMS 2019

Many of Genovis’ customers have submitted abstracts for ASMS 2019 in Atlanta, in which the SmartEnzymes have been used. Below is a digested guide to these abstracts so that you can plan your days at ASMS.

To read the full poster abstracts, visit the Online Planner for ASMS, and paste the abstract titel in the search field.

 

Sunday, June 2

New Enzymatic Workflows for Analysis of O-Glycosylated Biopharmaceuticals

Andreas Nägeli

Thermo Fisher Scientific User Meeting

Pharma/BioPharma Breakout Session 9:30 AM – 12.15 PM

Marquis Ballroom C

Thermo-user-meeting-blog

Monday, June 3

Improved middle-down characterization of antibodies using multiple ion activation techniques and Proton Transfer Reaction on a modified Orbitrap mass spectrometer 

Romain Huguet

MP 676, Poster Session 10.30 AM – 2.30 PM

FabRICATOR® and GingisKHAN®

 

Direct Determination of Antibody Chain Pairing by Top-Down Mass Spectrometry using Electron Capture Dissociation and Ultraviolet Photodissociation 

Jared Shaw

Room B302-305, Oral Presentation 3.50 PM – 4.10 PM

FabRICATOR® and GingisKHAN®

 

Unraveling a complex immunoprotein profile in multiple myeloma with middle-down de novo sequencing and native mass spectrometry 

Valerie J Winton

Room A411-412, Oral Presentation 9.30 AM – 9.50 AM

FabRICATOR®

 

Middle Down Approach for the Characterization of Monoclonal Antibodies After Ides Digestion and ETD Fragmentation

Colin Wynne

MP 782, Poster Session 10.30 AM – 2.30 PM

FabRICATOR®

 

Tuesday, June 4

Analysis of Zika Viral Polyprotein N- and O-glycosylation Using a Novel Lectin-chemoenzymatic Enrichment 

Shuang Yang

TP 655, Poster Session 10.30 AM – 2.30 PM

OpeRATOR® and GlycOCATCH™

 

Comprehensive Characterization of Antibody Drug Conjugates Enabled by Top-down and Middle-down Mass Spectrometry Strategies

Eli J Larson

TP 601, Poster Session 10.30 AM – 2.30 PM

GlycINATOR®FabRICATOR® and GingisKHAN®

 

Wednesday, June 5

Analysis of O-glycosylated Biopharmaceuticals using an O-glycan dependent Endoprotease and LC-MS

Andreas Nägeli

WP 334, Poster Session 10.30 AM – 2.30 PM

OpeRATOR® and OglyZOR®

 

MALDI-in-source decay FT-ICR MS for top-down and middle-down characterization of monoclonal antibodies 

Simone Nicolardi

WP 032, Poster Session 10.30 AM – 2.30 PM

FabRICATOR® and GingisKHAN®

 

Application of a label-free and domain-specific free thiol method in monoclonal antibody characterization

Yi Pu

WP 040, Poster Session 10.30 AM – 2.30 PM

FabALACTICA® and FabRICATOR®

 

Collision induced unfolding experiments to decipher the structural regions of a hybrid monoclonal antibody

Thomas Botzanowski

WP 481, Poster Session 10.30 AM – 2.30 PM

IgGZERO®

 

Thursday, June 6

Intact and Subunit Mass Analysis Using Native Ion Exchange Chromatography Coupled to an Orbitrap Mass Spectrometer 

Qian Liu

ThP 663, Poster Session 10.30 AM – 2.30 PM

FabRICATOR®

 

Ultra-Fast Analysis of Intact Proteins Using SPE- TOF 

Kevin McCann

ThP 149, Poster Session 10.30 AM – 2.30 PM

FabRICATOR®

 

Isomeric linkage determination of Sialic acid on O-glycopeptides using O-protease and LC-MS/MS 

Jieqiang Zhong

ThP 071, Poster Session 10.30 AM – 2.30 PM

OpeRATOR®

Free Thiols using FabRICATOR® and FabALACTICA®

In biopharmaceutical product development and manufacturing, free thiol content is one of the product quality attributes of interest as its presence could impact structure, stability and function of the product.

At Biogen, Yi Pu et al have optimized a label-free LC (UV) / MS method for free thiol quantification at a subunit level of IgG1 and IgG4. The new method, which is based on a method developed by Faid et al*, was compared to two conventional approaches, Ellman’s assay and peptide mapping.

It is very challenging to identify free thiol forms by mass spectrometry at the intact antibody level. By combining the highly specific proteolytic enzymes FabALACTICA (IgdE) and FabRICATOR (IdeS) the authors generated the subunits Fab, hinge and Fc/2, suited for confident mass determination. The subunits were subsequently separated on a polyphenyl reversed phase column in order to separate free thiol forms from their corresponding disulphide bond-linked form. A baseline or near baseline separation was obtained making it possible to calculate the free thiol content on each subunit.

The result of the quantification of free thiols from all three methods were comparable and showed similar trends even though the peptide mapping approach generally gave a higher free thiol content.

The authors conclude that compared to Ellman’s assay, the subunit approach is more sensitive, requires less sample and provides domain-specific information of the free thiol content. Compared to peptide mapping, the subunit method is faster, less labour intensive and lacks dependence on labelling efficiency. Finally, it demonstrated promise in the quantification of free thiols in a high throughput manner with domain specific information available.

The developed method has successfully been applied to several in-house IgG1 mAbs with different hydrophobicity and isoelectric points.

 

*V. Faid Y. Leblanc N. Bihoreau G. Chevreux Middle-up analysis of monoclonal antibodies after combined IgdE and IdeS hinge proteolysis: Investigation of free sulfhydryls, J. Pharm. Biomed. Anal. 149 (2018) 541-546, https://doi.org/10.1016/j.jpba.2017.11.046

 

For more information on FabRICATOR and FabALACTICA please visit the following pages:

The full text paper is available online:

The Digested Guide to ASMS 2018

 

Several researchers have submitted abstracts for ASMS 2018 in San Diego, in which Genovis’ SmartEnzymes have been used. Below is a selection of these abstracts.

To read the poster abstracts, visit the Online Planner for ASMS, and paste the abstract titel in the search field.

Monday June 4 

10:30AM-2:30PM: Poster Session

MP 045 – GlyCLICK

Development of NISTmAb-derived homogeneous antibody-drug conjugate (ADC) standards

Shanhua Lin1; Terry Zhang2; Brian Agnew3; Trina Mouchahoir4; John Schiel4
1Thermo Fisher Scientific, Sunnyvale, CA; 2Thermo Fisher Scientific, San Jose, CA; 3Thermo Fisher Scientific, Eugene, Oregon; 4NIST, Gaithersburg, MD

MP 046 – FabRICATOR

Discovery and confirmation of glucuronylation as a new acidic post-translational modification on therapeutic monoclonal antibodies

Yuetian Yan1; Anita Liu1; Shunhai Wang1; Thomas Daly1; Ning Li1
1Regeneron Pharmaceuticals, Tarrytown, NY

MP 049 – FabRICATOR

Ultrasensitive Characterization of Size and Charge Heterogeneity of Therapeutic Monoclonal Antibodies by Native Mass Spectrometry

Shunhai Wang1; Yuetian Yan1; Anita Liu1; Thomas Daly1; Ning Li1
1Regeneron Pharmaceuticals, Tarrytown, NY

MP 464 – FabALACTICA

Quantitative UPLC-MSE analysis of disulfide bonds and free sulfhydryls in monoclonal antibodies using IgdE protease assisted digestion

Jeroen de Keijzer1; Peter van Maurik1; Anja Boumeester1; Emile van Corven1; Gideon Oudgenoeg1
1Bioceros, Utrecht, Netherlands

 

Tuesday June 5 

10:30AM-2:30PM: Poster Session

TP 624 – FabRICATOR

Avoiding method induced heterogeneity in the analysis of heterogeneity of monoclonal antibodies using Mass Spectrometry after Single Site Proteolysis

Gideon Oudgenoeg1; Anja Boumeester2; Peter van Maurik2; Jeroen de Keijzer2; Emile van Corven2
1Bioceros, Utrecht, Netherlands; 2Bioceros, Utrecht, Netherlands

 

Wednesday June 6 

10:30AM-2:30PM: Poster Session

WP056 – FabRICATOR

Characterizing and Quantitating Therapeutic Antibody Multimer Degradation Using Affinity Capture Mass

Neha Srikumar1; Wenjing Li1; Robert Tchelepi1; Chen Gu1; Diego Ellerman1; Greg A Lazar1; Yichin Liu1John C. Tran1
1Genentech Inc., South San Francisco, CA

WP 327  – GingisKHAN

Comprehensive Domain-Specific [Fc vs. Fab] N-glycosylation Analysis of Therapeutic Proteins

Charles Nwosu1; Shuangqi Sally Liu2; Lei Wang2; May Zhu2; Anne Kowal2
1Takeda Pharmaceuticals International Co, Cambridge, MA; 2Takeda Pharmaceuticals International Co., Cambridge, MA

WP 340 – FabULOUS

Analysis of Fragmented Porcine Immunoglobulin G (IgG) by MALDI-MS and UPLC-ESI-MS

HELENE PERREAULT1; Claudia Nelson2
1University of manitoba, Winnipeg; 2University of Manitoba, Winnipeg, MB

WP 342 – OpeRATOR and GlycOCATCH

A Novel O-glycoprotease with applications in O-glycan Analysis using mass spectrometry

Rolf Lood1, 2; Maria Nordgren1; Fredrik Leo1; Stephan Björk1; Malin Mejáre1Fredrik Olsson1
1Genovis AB, Lund, Sweden; 2Department of Infectious Diseases, Lund University, Lund, Sweden

WP 350 – OpeRATOR

Deciphering complex o-glycosylation: solid-phase chemoenzymatic cleavage and enrichment

Shuang Yang1; Philip Onigman2; Jonathan Sjogren2; Wells W. Wu3; Rong-fong Shen3; John Cipollo1
1LBP, CBER, FDA, Silver Spring, MD; 2Genovis AB Inc., Boston, MA; 3FBR, CBER, FDA, Silver Spring, MD

WP 676 –FabRICATOR, GlycINATOR, IgGZERO

Monitoring Critical Quality Attributes: Core Fucosylation of N-glycans using an Integrated Subunit LC/MS Workflow Method

Nilini S Ranbaduge1; Henry Y Shion1; Ying Qing Yu1; Weibin Chen1
1Waters Corporation, Milford, MA

WP 678 – FabRICATOR

In-depth Characterization of the Heterogeneous Dimerization Interfaces of A Monoclonal Antibody: from Subdomain Level to Residue Level

Yuetian Yan1; Shunhai Wang1Thomas Daly1; Ning Li1
1Regeneron Pharmaceuticals, Tarrytown, NY

W 691 – OpeRATOR, OglyZOR and SialEXO

LC-MS Characterization of Complex Glycoproteins

Amber Peariso1; Jason X. Tang1
1Eli Lilly & Company, Indianapolis, IN

WP 692 – FabRICATOR

Rapid Identity Assays for mAb Development, Production Control and Release

Anja Resemann1; Waltraud Evers1Yue Ju2; Guillaume Tremintin2; Detlev Suckau1
1Bruker Daltonics, Bremen, Germany; 2Bruker Daltonics, Billerica, MA

 

Thursday June 7 

10:10AM-10:30PM: Oral Presentation

Oral Presentation – FabRICATOR

Classification of Plasma Cell Disorders by 21 Tesla FT-ICR Top-Down and Middle-Down MS/MS Analysis of Monoclonal Immunoglobulins in Human Serum

Lidong He1, 2; Lissa C Anderson2; David R Barnidge3; David L Murray4; Surendra Dasari4; Angela Dispenzieri4; Christopher L Hendrickson1, 2; Alan G Marshall1, 2
1Florida State University, Tallahassee, FL; 2National High Magnetic Field Laboratory, Tallahassee, FL; 3The Binding Site, Rochester, MN; 4Mayo Clinic, Rochester, MN

 

10:30AM-2:30PM: Poster Session

ThP 003 – FabRICATOR

LC-MS in Combination with Multiple Enzymatic Digestion for Sequence Variant Identification in Support of Cell Line Development

Renpeng Liu1; Lintao Wang1; Alexandru C. Lazar1
1ImmunoGen, Waltham, MA

ThP 017 – GingisKHAN

Consortium for Top-Down Proteomics Inter-laboratory Study for Characterizing Monoclonal Antibodies (mAbs) by Top-Down Mass Spectrometry

Kristina Srzentic1; Luca Fornelli1; Yury Tsybin2; Joseph Loo3; Jeffrey Agar4; Julia Chamot-Rooke5Paul Danis6; Ying Ge7; David Goodlett8; Neil Kelleher1; Ljiljana Pasa Tolic9; Lloyd Smith7; Timothy Toby1; Konstantin Nagornov2; JENNIFER BRODBELT10; Sylvester Greer10; Mathieu Dupré5; David Clarke11; Ziqing Lin7; Kim Haselmann12; Christopher Hendrickson13; Lidong He13; Donald Hunt14; Jared Shaw9; Wendy Sandoval15; Richa Sarin16; Detlev Suckau17; Yuri E.M. van der Burgt18; Norelle Wildburger19; Nicolas L. Young20; Alain Beck21; John Yates22; Jolene Diedric22; Sneha Chatterjee23; Frank Sobott24; Anton Kozhinov2; ALAN G. MARSHALL13; LISSA C. ANDERSON13; Natalia Gasilova25; Laure Menin25; Neil Quebbenamm3; Sung Hwan Yoon26; Josh Hinkle14; Simone Nicolardi18; Matthew V. Holt20; Yunqiu Chen16; Nicholas Schmitt4
1Northwestern University, Evanston, IL; 2Spectroswiss Sàrl, Lausanne, Switzerland; 3UCLA, Los Angeles, CA; 4Northeastern University, Boston, MA; 5Institute Pasteur, Paris, France; 6Eastwoods Consulting, Boylston, MA; 7University of Wisconsin, Madison, WI; 8University of Maryland, Baltimore, Baltimore; 9PNNL, Richland, WA; 10University of Texas at Austin, Austin, TX; 11Edinburgh University, Edinburgh, United Kingdom; 12Novo Nordisk, Malov, Denmark; 13National High Magnetic Field Laboratory, Tallahassee, FL; 14University of Virginia, Charlottesville, VA; 15Genentech, Inc., South San Francisco, CA; 16Biogen Inc, Cambridge, MA; 17Bruker Daltonik GmbH, Bremen, Germany; 18Leiden University Medical Centre, Leiden, Netherlands; 19Washington University, St. Louis, St. Louis, MO; 20Baylor College of Medicine, Houston, TX; 21Centre d’immunologie Pierre Fabre, Saint-Julien-en-Genevois, France; 22The Scripps Research Institute, La Jolla, CA; 23University of Antwerp, Antwerp, Belgium; 24University of Leeds, Leeds, United Kingdom; 25Ecole Polytechnique Fédérale de Lausanne, Ch-1015 Lausanne, Switzerland; 26University of Maryland, Baltimore, MD

ThP 028 – FabRICATOR

Top- and Middle-Down CE-ESI-MS Analysis of Intact mAbs Using the ZipChip Coupled to a Fusion Lumos ETD Mass Spectrometer

Tricia C. Ho1; Erik J. Soderblom1; Erin Redman2; Greg M. Waitt1; M. Arthur Moseley1
1Duke University School of Medicine, Proteomics and Metabolomics Shared Resource, Durham, NC; 2908 Devices, Inc., Carrboro, NC

ThP 419 – FabRICATOR

Quantitation of Misincorporations: Strategies and System Suitability

Kathleen Cornelius1; Olga Friese1; Mary Denton2; Jason Rouse2
1Pfizer, Inc, Chesterfield, MO; 2Pfizer Inc., Andover, MA

 

2:30 -2:50 PM: Oral Presentation

Ballroom 20D – FabRICATOR, GingisKHAN, GlycINATOR and IgGZERO

A Suite of Liquid Chromatography Strategies Coupled Online to Top-down High-resolution Mass Spectrometry for Comprehensive Analysis of Antibody Drug Conjugates

Bifan Chen1; Ziqing Lin1; Qingge Xu1; Cexiong Fu2; Qunying Zhang2; Ying Ge1
1University of Wisconsin-Madison, Madison, Wisconsin; 2Abbvie Inc., North Chicago, IL

Interview with Valegh Faid at LFB Biotechnologies in France

 

Unique enzymatic digestions in study of antibody disulphides

 

Valegh Faid and colleagues at LFB Biotechnologies in France have developed and published an assay to study antibody disulphide bonds using middle-up LC-MS (Faid et al., 2017). The combination of FabRICATOR® for digestion below the hinge and FabALACTICA™ for digestion above the hinge, generated three fragments from a human IgG1 antibody; the hinge peptide, Fab and Fc/2 fragments. These fragments were resolved using RP-HPLC and mass spectrometry and enabled analysis of antibody disulphide bridges and other quality attributes.

 

 

Interview with Valegh Faid, Scientist at LFB and first author of the paper:

 

Why are antibody disulphide bonds important?

 

Disulphide bonds are highly important because of their critical role in the stabilization of protein conformations. Breaking and/or scrambling of disulphide bond occur during manufacturing and storage of biotherapeutics which is a concern in terms of safety and efficacy. The monitoring of these product-derived impurities is mandatory during development operations in order to minimize these forms.

 

How did you come up with the idea to combine FabRICATOR (IdeS) and FabALACTICA (IgdE)?

 

We have been using IdeS for many years in order to cleave IgG’s below the hinge; following DTT reduction, more amenable fragments for RP-HPLC/MS analysis are generated as previously published by our laboratory (Chevreux et al., 2011). This middle-up analysis is fast and very informative regarding the protein sequence integrity and post-translational modifications. However, investigating the oxidative state of disulfide bridges is tricky and often involved a time-consuming peptide mapping in non-reducing conditions.

In this context, IgdE is an interesting enzyme that cleaves specifically IgGs above the hinge and without requiring reducing conditions as papain do. The combination of IdeS and IgdE in non-reducing conditions presents the advantage to generate specifically three fragments i.e. hinge, Fc/2 and Fab that are both easily separated by RP-HPLC and analysed by MS.

 

How does the new enzymatic assay compare to previous methods to study antibody disulphide bonds?

 

Peptide mapping in non-reducing condition is the gold standard to investigate disulphide bonding of biotherapeutics. However, data interpretation is time consuming even if dedicated software to improve the treatment of data has largely improved. Although being slightly less informative than peptide mapping, this combined IdeS/IgdE middle-up approach increases the throughput for the investigation of free thiols and disulphide scrambling. Considering that other CQAs can also be monitored in the same experiment, it should be more applicable to routine use in process optimization, formulation screening and stability studies.

 

Would the assay be used in a QC setting relying solely on liquid chromatography separation?

 

The analytical workflow is robust and requires mere handlings of the antibody samples. Once the identification of each peak of the chromatogram is confirmed by MS, quantitation based on the UV detection is a current practice. Such analytical configuration involving an HPLC and a UV detection is actually common in most of QC labs and thus easily and robustly implementable.

 

How are you implementing this assay at LFB Biotechnologies?

 

This assay is integrated in our portfolio of analytical approaches for the analysis of mAbs currently in development, for process optimisation, batch characterization and stability studies.

 

 

Read more about FabALACTICA and FabRICATOR.

 

References:

 

Chevreux, G. et al., 2011. Fast analysis of recombinant monoclonal antibodies using IdeS proteolytic digestion and electrospray mass spectrometry. Anal Biochem. 15;415(2): pp. 212-4.

 

Faid, V. et al., 2017. Middle-up analysis of monoclonal antibodies after combined IgdE and IdeS hinge proteolysis: Investigation of free sulfhydryls. Journal of Pharmaceutical and Biomedical Analysis, 149, pp.541–546.

 

 

Meet Our New Colleague Kevin Cook

May 17, 2018 | Genovis Team |

 

We are happy and proud to have you on board, Kevin. Welcome to Genovis!

 

Tell us a bit about yourself.
Even though a Focused Generalist does not make perfect sense, I think it fits since I have a broad life sciences discovery background with a specificity towards LC-MS and orthogonal technology.

 

If you were to describe yourself using only one word – what would that word be?

Curious.

 

Tell us a bit about your previous working life.

My on-the-job learning experiences, with small and large companies, started with DNA/RNA focused laboratory work, moved into LC-MS to support small molecule pharmacokinetics and drug metabolism, and has migrated into a focus on providing biologics sample preparation technology for LC-MS analysis.

 

What will your main focus be here at Genovis?

On behalf of the Genovis team, I have the pleasure of building relationships with scientists in the western US. I am excited to learn more about the challenges my colleagues in the industry are facing and how our team can help.

 

What do you believe will be the biggest opportunity in your new position as a Senior Application and Market Area Manager?

Genovis has built a customer centric reputation that shows in current products and formats, and how LC-MS scientists are utilizing these tools. Genovis has demonstrated the ability to rapidly respond to customer requests and I am looking forward to helping promote current products while building for the future through customer feedback and collaboratory efforts.

 

4 Quick Questions:

Coffee or tea? 

Coffee, lots of coffee…

 

Aerosmith or Depeche Mode?

At the moment, Eric Church.

 

Ice cream or candy?

Chocolate chip cookies.

 

Soccer or ice hockey?

American football though staying with the question, ice hockey – Chicago Blackhawks.

Study on Glycoform Heterogeneity using Enzymatic Digestion and Native Mass Spectrometry

In a study by Wohlschlager et al. (2018), FragIT™ kit was used to digest the Fc-fusion protein etanercept, and the resulting fragments were analyzed using high-resolution native mass spectrometry (MS). Native MS offers a higher spatial resolution at a lower charge state, enabling studies of glycan heterogeneity, and FragIT digestion reduces sample complexity, enabling a detailed annotation of glycoforms on complex compounds.

 

A detailed knowledge about structure and post-translational modifications (PTMs) is required for biopharmaceuticals to be approved for clinical use, and an important quality attribute that may affect both the efficacy and safety of biopharmaceuticals is glycosylation.

 

Etanercept is a highly glycosylated Fc-fusion protein that is used to treat autoimmune diseases such as rheumatoid arthritis, and it consists of the TNF-𝛼 receptor domain fused to the Fc domain of human IgG1. FragIT – an immobilized version of the FabRICATOR® (IdeS) enzyme – digests IgG from several species and subclasses at a specific site below the hinge region. The resulting fragments are easily purified using the Fc-specific affinity resin that, together with FragIT, comprises the FragIT kit.

 

In this study, the researchers analyzed the glycosylation of etanercept on both the intact level, and on the middle-down level after FragIT digestion. By combining native MS analysis with enzymatic remodelling of etanercept, a detailed annotation of glycoforms could be achieved and transferred from subunit to whole protein level.

 

The authors end by concluding:

Comprehensive information on glycoform heterogeneity, fast analysis with minimal sample preparation and product-characteristic fingerprints render our method highly attractive for the quality control of biologics as well as for comparability studies following changes in the manufacturing process.”(Wohlschalger et al. 2018).

 

Read more about FragIT and FabRICATOR

 

References:

 

Genovis Launches GlycOCATCH™

April 26, 2018 | Applications, Products |

Please welcome GlycOCATCH™ to the Genovis SmartEnzymes™ family!

 

GlycOCATCH is an enrichment resin for affinity purification of O-glycosylated proteins and peptides. The resin is designed to bind proteins and peptides carrying O-glycans with high affinity, and it is provided in convenient spin columns to allow easy-to-use O-glycoprotein enrichment.

 

The applications of GlycOCATCH involve glycomics, specific enrichment or removal of O-glycoproteins and peptides, studies of O-glycosylation in complex samples and characterization of biopharmaceuticals.

 

The GlycOCATCH product is available for purchasing today.

 

Read more about GlycOCATCH here

A New Assay to Study IgG Galactosylation in Serum

March 21, 2018 | Applications, Products |

IgGZERO-glycation

In a study by Vanderschaeghe et al. (2018), a new assay to measure IgG galactosylation in serum has been developed. The setup includes hydrolysis of IgG Fc glycans using the IgG-specific endoglycosidase IgGZERO® (EndoS).

 

A reduced level of IgG galactosylation in serum is a promising biomarker to evaluate the severity, determine the treatment and assess the efficacy of the treatment of autoimmune diseases such as systemic lupus erythematosus (SLE), rheumatoid arthritis (RA) and Crohn’s disease.

 

Traditionally, it has been difficult to study IgG galactosylation in serum because of the requirement to purify the antibodies, a procedure that is both complex and time-consuming. However, Vanderschaeghe et al. demonstrate a new assay where IgGZERO is used to efficiently hydrolyze serum IgG Fc glycans before analyzing galactosylation on high-throughput DNA sequencers. IgGZERO works on natively folded IgG, meaning that the assay can be performed on complex serum without first needing to purify the IgG, a feature that renders the assay both fast and simple.

 

The authors conclude by describing their new IgGZERO-based assay:

“… an important breakthrough towards the clinical implementation of the proposed biomarker” (Vanderschaeghe et al. 2018).

 

Read more about IgGZERO

 

Find the full text of the paper here:

Vanderschaeghe et al., 2018. Clinical assay for direct assessment of IgG galactosylation in serum using endoglycosidase S. BioRxiv.