Polyacrylamide Gel Electrophoresis: An Analytical Technique to Detect Heparan Sulfates of Varying Chain Lengths Isolated from Mouse Lung Tissue

Published: April 30, 2023

Abstract

Source: LaRiviere, W. B. et al., Detection of Glycosaminoglycans by Polyacrylamide Gel Electrophoresis and Silver Staining. J. Vis. Exp. (2021)

This video demonstrates the separation procedure of heparan sulfates isolated from mouse lung tissue using polyacrylamide gel electrophoresis.

Protocol

1. Polyacrylamide gel electrophoresis of isolated and purified glycosaminoglycans Prepare solutions necessary for polyacrylamide gel electrophoresis (PAGE) in advance (Table 1).      NOTE: Select the percent acrylamide of resolving gel solution depending on the size of the glycosaminoglycans expected to be in the sample. 15% is recommended for resolving larger fragments (greater than 30 disaccharide subunits in length); 22% for…

Representative Results

Table 1: Solutions required for polyacrylamide gel electrophoresis of purified glycosaminoglycans. All solutions must be filtered (0.22 µm) before use. NOTE: All solutions must be filtered (0.22μm) before use. Solution Recipe Composition …

Offenlegungen

The authors have nothing to disclose.

Materials

Ammonium persulfate (solid) ThermoFisher Scientific BP179-25 Electrophoresis grade
Barnstead GenPure Pro Water Purification System ThermoFisher Scientific 10-451-217PKG Any water deionizing/ purification system is an acceptable substitute
Boric acid (solid) ThermoFisher Scientific A73-500 Molecular biology grade
Bromphenol blue (solid) ThermoFisher Scientific B392-5
Criterion empty cassette for PAGE (1.0mm thick, 12+2 wells) Bio-Rad 3459901 Any 1.0mm thick PAGE casting cassette system will suffice
Criterion PAGE Cell system (cell and power supply) Bio-Rad 1656019 Any comparable vertical gel PAGE system will work)
EDTA disodium salt (solid) ThermoFisher Scientific 02-002-786 Molecular biology grade
Glycine (solid) ThermoFisher Scientific G48-500 Electrophoresis grade
Hydrochloric acid (liquid) ThermoFisher Scientific A466-250
N,N'-methylene-bis-acrylamide (solid) ThermoFisher Scientific BP171-25 Electrophoresis grade
Phenol red (solid) ThermoFisher Scientific P74-10 Free acid
Sucrose (solid) ThermoFisher Scientific BP220-1 Molecular biology grade
TEMED (N,N,N',N'-tetramethylenediamine) ThermoFisher Scientific BP150-20 Electrophoresis grade
Tris base (solid) ThermoFisher Scientific BP152-500 Molecular biology grade
Vacuum filter unit, single use, 0.22uM pore PES, 500mL volume ThermoFisher Scientific 569-0020 Alternative volumes and filter materials acceptable
Acrylamide (solid) ThermoFisher Scientific BP170-100 Electrophoresis grade
Heparin derived decasaccharide (dp10) Galen scientific HO10
Heparin derived hexasaccharde (dp6) Galen scientific HO06
Heparin derived oligosaccharide (dp20) Galen scientific HO20

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Diesen Artikel zitieren
Polyacrylamide Gel Electrophoresis: An Analytical Technique to Detect Heparan Sulfates of Varying Chain Lengths Isolated from Mouse Lung Tissue. J. Vis. Exp. (Pending Publication), e21106, doi: (2023).

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