Waters GlycoWorks Single Use Sample Preparation Kit - Manual (Page 2)

GlycoWorks Single Use Sample Preparation Kit 2
[ CARE AND USE MANUAL ]
Table 1: Kit Components
Kit Component Purpose
GlycoWorks
HILIC 1 cc
Cartridge
Extract and clean up of free or FLR labeled glycans
■ Capture of glycans
■ Removing contaminants like salt and
detergents from hydrophilic analytes, such
as carbohydrates, prior to MS
■ Removing excess labeling reagent
■ The 10 mg sorbent was test to have maximum
binding capacity for 400 µg glycans
GlycoWorks
Glycoprotein
Control
Standard Unlabeled IgG (100 µg/lyophilized) to
use as a control for process validation and
troubleshooting
RapiGest SF An enzyme friendly detergent used to denature
the glycoproteins prior to deglycosylation
reaction. For stability purposes, there is enough
RapiGest included for initial use of the kit with
5 samples, more may need to be ordered for
processing all 10 samples.
Manifold
Waste Tray For collection of non-releasing glycans steps.
This is disposable but can be cleaned and used
several times before acquiring a new one as long
as it is rinsed out.
GlycoWorks
Reagent Kit Contains ready to use reagents for both
denaturation, reduction, and alkylation as well as
raw materials for FLR tagging.
II. STORAGE AND DISPOSAL OF CARTRIDGES
Cartridges stored in their original sealed pouch remain stable for
long periods. For long term storage of unused cartridges, keep
cartridges sealed and in a desiccator.
Note: Dispose of used cartridges safely in accordance with applicable
government or local regulations
III. ABBREVIATIONS
DMSO Dimethylsulfoxide IAM Iodoacetamide
DTT Dithiothreitol 2AB 2-aminobenzamide
IV. USING THE GLYCOWORKS SINGLE USE KIT
Below, are general guidelines for utilizing the GlycoWorks Single
Use Kit for complete extraction, purification, and concentration of
N-linked glycans validated using IgG. Also included is a check list of
accessory materials that will be needed to complete the protocol from
start to finish as well as tips and tricks throughout the instructions.
Table 2: Checklist of Additional Materials Needed Before You Begin
Material Description
Recommended
Suppliers
Check List:
Circle
Ammonium
bicarbonate Sigma Aldrich
Cat No. O9830
Yes No
Ammonium acetate
Sigma Aldrich
Cat No. 73594 FLUKA
Yes No
Formic acid (to make
1% [v/v] formic acid,
prepared in 50%
acetonitrile)
Sigma Aldrich
Cat No. F0507
Yes No
15,000 units
(500,000 units/mL)
PNGase F*
New England Biolabs
Cat. No. P0704S
Yes No
Acetonitrile
Sigma Aldrich
Cat No. 360457
Yes No
25 mM ammonium
bicarbonate in water,
pH 8 -9
See Above Yes No
100 mM ammonium
acetate in 5% [v/v]
acetonitrile, pH 7
See Above Yes No
85% [v/v] acetonitrile See Above Yes No
2-AB** Sigma P/N A89804 Yes No
SPE vacuum pump
115, 60 Hz:
Waters P/N 176002986
Yes No
Centrifugal vacuum
evaporator
N/A Yes No
Heat block N/A Yes No
Fisherbrand Premium
1.5 mL MCT graduated,
RNase/DNase free
Fisher Scientific Cat No.
05-408-129
Yes No
* PNGase F comparison table: https://www.neb.com/tools-and-resources/
usage-guidelines/glycobiology-unit-conversion-chart
**Inclusive Kits are also offered by ProZyme, Ludger, and Sigma covered
under US Patent No. 5,747,347 and its foreign equivalents.