Cy5 Labeled Heparan Sulfate Proteoglycan (Cy5-HSPG)

Catalog #: GL402 Datasheet / COA / SDS

Catalog #	Availability	Size / Price	Qty
GL402-100

Cy5 Labeled Heparan Sulfate Proteoglycan (Cy5-HSPG).

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Cy5 Labeled Heparan Sulfate Proteoglycan (Cy5-HSPG) Summary


Formulation	Supplied in 25 mM Tris, 150 mM NaCl, pH 7.5
Stability & Storage	Store at < -20 °C. Good for 6 months from date of receipt.

Applications

Used as a substrate to study activity of various heparanases and heparinases.
Used to study protein heparan sulfate interactions.

Key Features:

Excitation at 649 nm and emission at 671 nm.
The fluorescent dye Cy5 is conjugated to the non-reducing end terminal GlcNAc residue.
Heparan sulfate can be released by digestion with heparanase or heparinase II, heparinase III.
Can be separated on 15%-17% SDS-PAGE and directly visualized.

For Details:

Wu et al., (2021) Glycobiology 31:1435-1443

Related Reagents

Click Chemistry

Enzymes and detection reagents

Recombinant Human Active Heparanase/HPSE Protein (7570-GH)
Recombinant Mouse Active Heparanase/HPSE Protein (9788-GH)
Recombinant Human Exostosin 1/2 Heterodimer Protein (8567-GT)

Specifications

Storage

Store the unopened product at -70 °C. Use a manual defrost freezer and avoid repeated freeze-thaw cycles. Do not use past expiration date.

Product Datasheets

Product Datasheet

COA

SDS

Scientific Data

N/A

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Cy5 Labeled Heparan Sulfate Proteoglycan (Cy5-HSPG). This substrate can be used to study the activity of various heparanases and heparinases. This can also be used to study protein heparan sulfate interactions. The fluorescent dye Cy5 is conjugated to the non-reducing end terminal GlcNAc residue.

SDS-PAGE

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Fluorescent gel mobility shift caused by Recombinant Mouse Heparanase. Lane 1-4 contained Cy5 Labeled Heparan Sulfate Proteoglycan (Cy5-HSPG) (Catalog # GL402) digested with increasing amounts of Recombinant Mouse Active Heparanase/HSPE Protein (9788-GH). Lane 5 contained Cy5-HSPG only. Lane 6, Cy5-HSPG was digested with Recombinant P. heparinus Heparinase III Protein (6145-GH). Lane 7, Cy5-HSPG was digested with Recombinant P. heparinus Heparinase II Protein (6336-GH).

Background: Heparan Sulfate Proteoglycan/HSPG

Long Name

Heparan Sulfate Proteoglycan

Entrez Gene IDs

3339 (Human); 15530 (Mouse)

Alternate Names

basement membrane-specific heparan sulfate proteoglycan core protein; endorepellin (domain V region); heparan sulfate proteoglycan 2; Heparan Sulfate Proteoglycan; HSPG; perlecan proteoglycan; perlecan; PLCSchwartz-Jampel syndrome 1 (chondrodystrophic myotonia); PRCAN; SJA; SJS; SJS1

Assay Procedure

Sample Assay Protocol for using Cy5 Labeled Heparan Sulfate Proteoglycan (Cy5-HSPG) as substrate for Recombinant Mouse Heparanase Assay

Protocols are guidelines. Parameters need to be optimized by end users. Suggested input of Cy5-HSPG in an assay separated on SDS-PAGE is 2 pmol.

Materials

Assay Buffer: 50 mM NaOAc, pH 5.0
Recombinant Mouse Active Heparanase/HPSE Protein (rmHPSE) (Catalog # 9788-GH)
Cy5 Labeled Heparan Sulfate Proteoglycan (Cy5-HSPG) (R&D Systems®, Catalog # GL402)
15% SDS-PAGE
6X Gel Loading Dye
A fluorescent imager

Assay

Dilute rmHPSE to 100 ng/µL and Cy5-HSPG to 0.4 µM with Assay Buffer.
Add 5 µL of rmHPSE, 5 µL of Cy5-HSPG, and 10 µL of Assay Buffer to a tube.
Prepare a negative control by mixing 5 µL of Cy5-HSPG and 15 µL of Assay Buffer in another tube.
Incubate the reaction and control at 37^o C for 60 minutes.
Add 5 µL of 6X Gel Loading Dye to each tube.
Load 12.5 µL of each tube to a 15% SDS-PAGE gel and run the gel at 250 constant voltage until the dye front reaches the end of the gel.
Expose the gel for 10 seconds on a fluorescent imager.
Determine change in fluorescent signal with Alphaview or any other densitometry software.