Cy5-Fuc Labeled Glycan G2f Summary
Key Benefits
Learn more about Fluorescent Glycan Labeling and Detection
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Formula |
Fuc1Gal2Man3GlcNAc4Cy5 |
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Molecular Weight |
2616.44 Da |
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Formulation |
Supplied in 20 mM Tris, pH 8.0 |
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Stability & Storage |
Store at < -20 °C. Good for 6 months from date of receipt. |
Applications
- Used as a ligand for lectins and to study glycan protein interaction.
- Used as a substrate for various glycosidases and glycosyltransferases.
- Used as a scaffold for the synthesis of various glycan epitopes such as sialyl Lewis X structures.
Key Features and Benefits:
- Excitation at 649 nm and emission at 671 nm.
- The fluorescent dye Cy5 is conjugated to the C6 position of the core-6 fucose.
- Can be separated on 15%-17% SDS-PAGE and directly visualized as a single band under a fluorescent imager.
- Linear response range for Cy5-labeled glycans can be from 10 fmol to 100 pmol, depending on the sensitivity of detection.
- Store the unopened product at < -20 °C. Good for 6 months from date of receipt.
For Details:
- Wu et al., (2019) Glycobiology 29:750-754
- Wu et al., (2020) Glycobiology 30:970-980
- A Simple SDS-PAGE-based Technique for Probing Terminal Glycans
Related Reagents
Click Chemistry
- GDP-Cy5-Fucose (ES301)
- CMP-Cy5-Sialic Acid (ES302)
- GDP-Cy3-Fucose (ES401)
- CMP-Cy3-Sialic Acid (ES402)
Enzymes and detection reagents
Data Examples
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Recombinant Sp Endo-S2 assay using Cy5-G2f (Catalog # GL302) as a substrate. Cy5-G2f (0.2 pmol per reaction) was digested by a serial dilution of Endo-S2 for 60 minutes at 37oC. The digestions were then separated on a 17% polyacrylamide gel and imaged with TCE staining (top panel) and a fluorescent imager (lower panel). Endo-S2 digestion on Cy5-G2f resulted in two products: unlabeled product 1 and labeled product 2. Product 2 is visible with 25 ng of Endo-S2 and achieves the maximal level at 100 ng of the enzyme. |
Specifications
Product Datasheets
Scientific Data
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Cy5-Fuc Labeled glycan G2f This substrate can be used as ligand for lectins and to study various glycosidases and glycosyltransferases. This can also be used as a scaffold for the synthesis of various glycan epitopes such as sialyl Lewis X structures. The fluorescent dye Cy5 is conjugated to the C6 position of the core-6 fucose.
Background: Glycan G2f
Assay Procedure
Sample Assay Protocol of using Cy5-Fuc Labeled Glycan G2f/Cy5-G2f as substrate for recombinant Sp. Endo-S2 assay
Protocols are guidelines. Parameters need to be optimized by end users. Suggested input of a Cy5-labeled glycan in an assay separated on SDS-PAGE is from 0.01 pmol to 1 pmol.
Materials
- Assay Buffer: 50 mM MES, pH 6.0.
- Recombinant Sp. Endo-S2
- Cy5-Fuc Labeled Glycan G2f (Cy5-G2f) (Catalog # GL302)
- 6X gel loading dye
- A fluorescent imager
Assay
- Dilute rSp. Endo-S2 to 10 ng/µL in Assay Buffer.
- Dilute Cy5-G2f to 0.02 µM in Assay Buffer.
- Mix 10 µL of the diluted rSp.Endo-S2 and 10 µL of the Cy5-G2f in a centrifuge tube.
- Prepare a negative control by mixing 10 µL of the Cy5-G2f with 10 µL of Assay Buffer.
- Incubate the reaction and control at 37o C for 60 minutes.
- Stop the reactions and controls by adding 5 uL of 6X gel loading dye to each tube.
- Load 12.5 µL each of the above samples per well on a 15% SDS-PAGE and run at 10 volts/cm until the dye front is more than two thirds of the gel.
- Image the gel with a fluorescent imager for 10 seconds.
- Determine percent digestion with Alphaview or any other densitometry software.
Final Assay Conditions Per Reaction
rSp. Endo-S2: 100 ng
Cy5-G2f: 0.2 pmol
FAQs
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