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Thermo Scientific™ GlycoLink™ IP Kit

Catalog No. PI88943
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25 reactions kit
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PI88943 25 reactions kit
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Catalog No. PI88943 Supplier Thermo Scientific™ Supplier No. 88943
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Description

Immobilize polyclonal antibodies or other glycoproteins onto agarose beads for immunoprecipitation or pull-down assays via periodate-oxidized sugars.

The Thermo Scientific GlycoLink IP Kit provides components for effective immunoprecipitation (IP and co-IP) based on small-scale, covalent, affinity-resin immobilization of antibodies and other glycoproteins via oxidized sugar groups.

The GlycoLink Kit uses hydrazide-activated UltraLink Resin to immobilize and perform 25 IP or Co-IP assays with different antibodies or glycoprotein ligands. The method is ideal for polyclonal antibodies, which typically have abundant carbohydrates (glycosylation) on their Fc portions. Monoclonal antibodies that contain adequate carbohydrates are also effective. Antibodies immobilized by this hydrazide-to-carbohydrate method have unobstructed antigen-binding sites and optimal purification capability. The prepared glycoprotein or antibody columns can be regenerated and reused at least five times for immunoprecipitation, co-immunoprecipitation or pull-down affinity-purification without significant loss in binding capacity.

Highlights:

  • Efficient immobilization – couple 2 to 10μg of oxidized antibody or other glycoprotein to 20μL of resin in 2 hours or less

  • Stable immobilization – resonance structure of the hydrazone bonds are sufficiently stable so the antibody remains coupled during elution

  • Preserves binding function – immobilizes IgG via the Fc region, thereby keeping both antigen binding sites available for capturing target

  • Versatile applications – immobilize any molecule that contains oxidizable sugars, including glycoproteins for use in purifying protein interaction binding partners

Includes:

Kit contains resin, buffers and reagents for immobilization; desalting columns; IP lysis/wash and elution buffers for immunoprecipitation; microcentrifuge spin columns and collection tubes; and loading buffer for SDS-PAGE

Recommended for:

Immunoprecipitation followed by electrophoresis and excision of the protein for mass spectrometry or sequencing; Immunoprecipitation and SDS-PAGE analysis of a protein with a molecular weight similar to the antibody's heavy or light chain; Identification of binding partners of immobilized glycoprotein

Co-immunoprecipitation for determining protein-protein interactions

Specifications

Capacity (Metric) 2 to 10 μg Oxidized Antibody or Glycoprotein Coupling to 20 μL Resin
Content And Storage Sufficient For: 25 IP/Co-IP reactions, each using 20 μL antibody-coupled resin
• UltraLink Hydrazide Resin, 0.5 mL
• GlycoLink Coupling Buffer, 50 mL
• Aniline, 0.2 mL
• Sodium meta-periodate, 0.5 g
• Wash Buffer, 60 mL
• Zeba Spin Desalting Columns,0.5 mL, 25 columns
• IP Lysis/Wash Buffer, 50 mL
• PBS Pack (makes 500 mL), 1 pack
• Elution Buffer, pH 2.8, 50 mL
• Lane Marker Sample Buffer, Non-reducing, (5X), 5 mL
• Pierce Spin Columns-Screw Caps, 25 columns
• Collection Tubes, 2 mL, 100 tubes

Store at 4°C.
Assay Affinity Purification
For Use With (Equipment) Microcentrifuge
Product Line GlycoLink
Quantity 25 reactions kit
For Use With (Application) Immunoprecipitation (IP)
Target Not Target-Specific
Type Immunoprecipitation/Co-Immunoprecipitation Kit
Sufficient For 25 Reactions
Format Kit
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Frequently Asked Questions (FAQs)

What is the difference between GlycoLink and CarboxyLink Gels?

GlycoLink gel is hydrazide-activated crosslinked beaded agarose, and it is useful for coupling glycoproteins via aldehydes formed from their sugars by sodium meta-periodate oxidation. Reaction of aldehydes with hydrazide- activated resin is catalyzed by aniline resulting in >90% coupling in 4 hours or less. CarboxyLink gel on the other hand is crosslinked beaded agarose activated with diaminodipropylamine (DADPA) and is useful for immobilizing carboxyl- containing biomolecules after EDC activation. Actually, both resins can be used with EDC to couple ligands via carboxylic acids.

Note: Both immobilization chemistries are available on UltraLink Support as UltraLink Hydrazide and UltraLink DADPA respectively.
Will Glycolink Gel work with chicken IgY?

Yes, Chicken IgY can be immobilized using GlycoLink Gel, due to the high level of glycosylation of the IgY.
Will GlycoLink Gel work well for conjugating all types of antibodies?

No. Monoclonal antibodies are typically under-glycosylated, and coupling efficiency to CarboLink Gel may be low. However, many monoclonal antibodies can be immobilized using GlycoLink Gel. Try the Pierce Glycoprotein Carbohydrate Estimation Kit (Cat. No. 23260) to determine if your monoclonal antibody is glycosylated.
What are the benefits of using a GlycoLink Coupling Resin?

The benefits include high-capacity immobilization of 1-10 mg of oxidized antibody or other glycoprotein per milliliter of resin. Immobilization is fast (in as few as 30 mins) and can achieve at least 90% coupling of most glycoproteins in less than 4 hours. The hydrazide-activated UltraLink resin conjugates only to purified glycoproteins whose sugar groups have been gently oxidized with periodate.

No stabilization step is required when using this resin, and antibody function is preserved as IgGs are immobilized via the Fc region, keeping both antigen binding sites available for capturing targets.

For Research Use Only. Not for use in diagnostic procedures.