Waters GlycoWorks High-throughput Sample Preparation Kit - Manual

GlycoWorks High-throughput Sample Preparation Kit 1
[ CARE AND USE MANUAL ]
LC-FLR
LC-FLR-MS
I. INTRODUCTION
This document provides information regarding the general care and
use of the Waters GlycoWorks™ High-throughput Sample Preparation
Kit for release of N-glycans. This protocol can be used as a guideline
and is validated using IgG but can also be used and optimized for
other N-linked glycoproteins. This sample preparation offers an
inclusive protocol that allows the user to source many of the critical
components needed to perform the preparation from one vendor.
The 96-well plate is good for 48 analyses.
For expected results using the protocol with the control standard,
please see Reference 1.
The guide provides the following:
■
An instruction template for new users that highlights the use of a
control standard (included in the kit) for first time validation as
well as subsequent troubleshooting
■
Protocol for LC/FLR and LC/FLR/MS
■
Tips and tricks throughout each section to help aid in the success
■ An online training video is also available (see Reference 3)
GlycoWorks High-throughput Sample Preparation Kit
CONTENTS
I. INTRODUCTION
II. STORAGE AND DISPOSAL OF PLATES
III. ABBREVIATIONS
IV. USING THE GLYCOWORKS HIGH-THROUGHPUT KIT
V. PREPARING LABELED GLYCANS FOR HILIC-FLR WITH
AN ACQUITY UPLC BEH GLYCAN COLUMN
VI. REFERENCES
VII. ORDERING INFORMATION
Figure 1: Sample Preparation from Glycoprotein to Labeled Glycans
Glycoproteins
FLR Labeling Reaction of Glycans
3
FLR Labeled Glycans
MALDI MS
Denaturation and Deglycosylation
RapiGest™ (if necessary)
DTT Reduction (if necessary),
IAM Alkylation (if necessary)
Deglycosylation by PNGase F
1
Extraction of Released Glycans
GlycoWorks HILIC µElution Plate
2
Excess Labeling Reagent Removal
GlycoWorks HILIC µElution Plate
4
General Guideline of Sample Preparation from Glycoprotein to
enrich FLR Labeled Glycans Using Reductive Amination Reaction.
If using MALDI please refer to Product Solution, Literature Code 720004660EN