Brachyspira hyodysenteriae Infection Regulates Mucin Glycosylation Synthesis Inducing an Increased Expression of Core-2 O-Glycans in Porcine Colon.

Responsible:

Vignesh Venkatakrishnan . Sahlgrenska Academy, University of Gothenburg . vignesh.venkatakrishnan@gu.se

Sample preparation

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1.Sample Origin

General information:
O-glycans released from soluble and insoluble pig mucins isolateed from healthy and Brachyspira hyodysenteriae infected colon tissues

Growth/harvest conditions for recombinantly produced material:
NA

Treatments and/or storage conditions:
Soluble and insoluble mucins were purified from pig colon tissues using isopycnic gradient centrifugation. O-glycans were released chemically using reductive beta-elimination

For commercial material, vendor and applicable item information:
NA

Generation of chemically derived material:
NA

2. Sample Processing

Purification steps:
Released O-glycans were desalted using strong cation exchange Dowex resin packed as a microcolumn in top of hydrphobic C18 material

2. Defined Sample

Sample name:
O-linked oligosaccharides

HPLC

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1. General settings

Instrument: Thermo Fisher 1100 Series

Manufacturer url: NA

Injector: NA

Injector settings: NA

2. Column

Manufacturer: In-house

Model: Custom made Hypercarb

Type of Chromatography: Liquid chromatography

Type of Material: Porous graphitized carbon

Column diameter: 0.25

Column length: 100

Particle size: 5

Manufacturer url: NA

3. Method run

Temperature: NA

Solvent a: 10mM ammonium bicarbonate

Solvent b: 10mM ammonium bicarbonate in 80% Acetonitrile

Other solvent: NA

Flow rate: 250 nL/min

Flow gradient: NA

Run time: 78 minutes

Phase: Normal

MS

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1. General features

Date stamp: Tue Aug 04 UTC 2015

Instrument: Thermo Fisher LTQ Linear Ion Trap ESI

Customizations from the manufacturer's specification:
NA

ion_mode: -

2. Ion sources

(a) Electrospray ionisation - ESI

Sprayer type : Liquid chromatography

Interface name: NA

Other data for the Interface: NA

Sprayer name: need to check

Other data for the sprayer: NA

Relevant voltages: NA

Degree of prompt fragmentation evaluated: No

In-source dissociation performed: No

Other parameters if discriminant for the experiment: NA

(b) MALDI

Plate composition (or type): NA

Matrix composition: NA

Deposition technique: NA

Relevant voltages: NA

Degree of prompt fragmentation evaluated: NA

PSD (or LID/ISD) summary: NA

Operation with or without delayed extraction: NA

Laser type: NA

Other laser related parameters: NA

url: NA

3. Ion transfer optics

Hardware options: NA

3. Post-source componentry

Collision-Induced Dissociation - CID

CID. Gas composition: NA

CID. Gas pressure: NA

CID. Collision energy: NA

Electron Transfer Dissociation - ETD

ETD. Reagent gas: NA

ETD. Pressure: NA

ETD. Reaction time: NA

ETD. Reagent atoms: NA

Electron Capture Dissociation - ECD

ECD. Emitter type: NA

ECD. Voltage: NA

ECD. Current: NA

TOF drift tube

TOF. Reflectron status: NA

Ion Trap

Ion trap. Final MS stage achieved: NA

Ion mobility

Ion mobility. Gas type : NA

Ion mobility. Pressure: NA

Ion mobility. Instrument parameters: NA

FT-ICR

Peak selection: NA

FT-ICR. Pulse: NA

FT-ICR. Width: NA

FT-ICR. IR: Decay time: NA

FT-ICR. IR: NA

FT-ICR. Other parameters: NA

Detectors

Detector type: NA