Newly designed polyacrylamide/dextran gels for electrophoresis protein separation: synthesis and characterization

Abstract

Newly designed gels for electrophoresis protein separation were synthesized from acrylamide, N,N -methylenebis (acrylamide) and dextran mixtures. Radical polymerization was initiated by ammonium persulfate and N,N,N ,Ntetramethylethylenediamine. The time dependence of absorbance during polymerization was monitored using UV-visible spectroscopy. The exothermic polymerization process exhibited a sharp rise of temperature reminiscent of the Trommsdorff effect. The swelling kinetics of the synthesized gels was examined in deionized water and buffer solutions. One of the challenges was to find an alternative to commercial products, sold as mixtures with no detailed chemical contents, commonly used in sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) for protein separation. For this reason, a systematic comparison was made of the properties of one of the most commonly used commercial gels, Duracryl  from Genomics Solution Inc., and those of the synthesized polyacrylamide/dextran gels.

Proteome Analysis of Potato Starch Reveals the Presence of New Starch Metabolic Proteins as Well as Multiple Protease Inhibitors

“…A stock solution of acrylamide (50 mL) was prepared using 14.6 g acrylamide and 390 mg N , N , N ′, N ′ methylene-bis-acrylamide as a cross-linking agent. 330 mg of Dextran (MW 500 000) was also incorporated to the previous solution in order to improve gel separation according to the protocol developed in ( Ainseba-Chirani et al, 2011 ). The polyacrylamide concentrations were 4% and 7 or 10% in stacking (0.5 M Tris-Base, 14 mM SDS, pH 6.8) and running (1.1 M Tris-Base, 0.4 M Tris-HCl and 20 mM SDS, pH 8.8) gels, respectively.…”

Proteomics applied to transfusion plasma: the beginning of the story

“…It yields maps of proteins initially present in the sample showing post‐translational modifications. In spite of the wide use of polyacrylamide gels, matrix optimizations have been recently reported tending to improve the resolution of bands/spots [89], to allow the separation of hydrophobic proteins [90] and to facilitate the in‐gel digestion [91]. In Blue Native PAGE [92, 93], proteins, especially membrane proteins and protein supercomplexes, are separated at a neutral pH using an anionic detergent (Coomassie Blue) associated to a charge‐shifting agent 6‐aminocaproic acid, to enhance protein solubilization.…”

A mechanically strengthened polyacrylamide gel matrix fully compatible with electrophoresis of proteins and nucleic acids

“…The addition of preformed linear chains of polyacrylamide to aqueous acrylamide/bisacrylamide solutions has also been shown to strengthen gels through the formation of laterally bundled polymer chains linked by hydrogen bonds within the gel without affecting pore size . However, the turbidity of gels upon polymerization, high‐molecular weight band distortion and staining artefacts were also reported . The opportunity remains to develop mechanically strengthened low cost matrices for PAGE separation of biomolecules, whilst maintaining the existing electrophoretic properties.…”