LudgerTag Permethylation of Glycans

Analysis of glycans by MALDI-MS provides mass composition data and is faster than UHPLC techniques (on average one sample can take as little as 1 minute for data acquisition), but sialylated glycans are usually more difficult to analyse because the sialic acid moieties are labile. Permethylation is a popular technique for the derivatisation of carbohydrates for MALDI-MS detection, as it enhances ionization efficiency, stabilizes the sialic acids and aids linkage analysis studies. However, conventional techniques used for permethylation are slow, labour intensive and therefore not suited to high throughput sample analysis.

We have developed a simple to use LudgerTag kit for permethylating glycans, LT-PERMET-VP96. The kit can be used to process 1-96 samples using either a manual method or an automated method that has been adapted to a liquid handling robot. It is suitable for N- or O-glycans and once permethylated, they can be analysed by MALDI-TOF-MS or LC-MS.

We have validated this technology and demonstrated that glycoprofiling data is reliable and comparable to the gold standard HILIC UHPLC method. This technology has been used for a range of high throughput studies at Ludger (and using the automated permethylation sample preparation time is under 5.5 hours for 96 samples).

Product/ordering information:

N-glycans *:

  • E-PNG01
  • PNGase F, sufficient for 60 samples (depending on protocol)
  • LC-PERMET-96
  • Enrichment plate, for up to 96 samples
  • LT-PERMET-96
  • Permethylation kit, for up to 96 samples *This product has been discontinued
  • LT-PERMET-VP96
  • Permethylation kit, for up to 96 samples (without methyl iodide)

*Note: although hydrazinolysis can also be used for N glycan release, we recommend PNGase F release for the workflow

O-glycans:

  • LL-HYDRAZ-A2
  • Hydrazinolysis kit, for up to 12 samples
  • LC-CEX-A6
  • CEX cartridges, pack of 6, for 6 samples
  • LT-PERMET-96
  • Permethylation kit, for up to 96 samples *This product has been discontinued
  • LT-PERMET-VP96
  • Permethylation kit, for up to 96 samples (without methyl iodide)

Standards:

  • CPM-C13-IGG-01
  • IgG Heavy (13C) Permethylated Glycan Library, sufficient for approx 20 MS runs
  • CPM-IGG-01
  • IgG Permethylated Glycan Library, sufficient for approx 20 MS runs

Posters & Publications

Ludger Publication - Permethylation - Analytical Chemistry

Automated High-Throughput Permethylation for Glycosylation Analysis of Biologics Using MALDI-TOF-MS

Shubhakar A, Kozak RP, Reiding KR, Royle L, Spencer DI, Fernandes DL, Wuhrer M.
Analytical Chemistry. 2016 Sep 6. 88(17):8562-9. doi: 10.1021/acs.analchem.6b01639. Epub 2016 Aug 15.

Ludger Sofarchi 2016 poster

Automated High-Throughput Permethylation for Glycosylation Analysis Using MALDI-TOF-MS

Shubhakar A, Spencer D, Wuhrer M, Fernandes DL
Presented at the 38th Annual Congress of Pharmacology Society of Chile
Castro, Chile. Nov 26-29th 2016

Ludger poster

Reliability of a semi-automated, high throughput (HT) MS based glycomics system for discovery of glycan biomarkers and Quality by Design (QbD) studies of glycoprotein therapeutics

Shubhakar A, Spencer DI, Wuhrer M, Fernandes DL
Presented at Gordon Research Conference
Lucca, Italy. March 2015

Ludger poster

Drug Glycosylation Analysis using Automated, High Throughput, Orthogonal Methods

Shubhakar A, Kozak RP, Badia-Tortosa C, Gardner RA, Royle L, Wuhrer M, Spencer DI, Fernandes DL
Presented at PEGS 2014: Glycosylation Analysis for Biopharmaceuticals Workshop
Boston, United States. May 2014

Ludger poster

Optimisation of drug glycosylation supported by an automated high throughput permethylation MALDI-MS method

Shubhakar A, Spencer DI, Kozak RP, Velez-Suberbie L, Bracewell DG, Fernandes DL, Wuhrer M
Presented at ICS: 27th International Carbohydrate Symposium
Bangalore, India. January 2014

Contacts