An efficient solubilization buffer for plant proteins focused in immobilized pH gradients

• Published in: Proteomics (Weinheim) Vol. 3; no. 7; pp. 1299 - 1302
• Main Authors: Méchin, Valérie, Consoli, Luciano, Le Guilloux, Martine, Damerval, Catherine
• Format: Journal Article
• Language: English
• Published: Weinheim WILEY-VCH Verlag 01-07-2003; WILEY‐VCH Verlag; Wiley-VCH Verlag
• Subjects: Biochemistry, Molecular Biology; Buffers; Detergents - pharmacology; Electrophoresis, Gel, Two-Dimensional - methods; Hydrogen-Ion Concentration; Immobilized pH gradients; Isoelectric Focusing - methods; Life Sciences; Plant Proteins - chemistry; Protein solubilization; Proteins - analysis; Proton-Motive Force; Reducing Agents - pharmacology; Silver Staining; Two-dimensional gel electrophoresis; GENETIQUE; BIOLOGIE MOLECULAIRE
• ISSN: 1615-9853; 1615-9861
• DOI: 10.1002/pmic.200300450

The solubilization of a large array of proteins before electrophoresis itself is a very critical point for proteomic analyses. We compared the efficiency of several different solubilization buffers. From this work, we defined a very efficient solubilization buffer, including two chaotropes, two reducing agents (R2), two detergents (D2), and two kinds of carrier ampholytes in combination. This so‐called R2D2 buffer (5 M urea, 2 M thiourea, 2% 3‐[(3‐cholamidopropyl) dimethyl‐ammonio]‐1‐propane‐sulfonate, 2% N‐decyl‐N,N‐dimethyl‐3‐ammonio‐1‐propane‐sulfonate, 20 mM dithiothreitol, 5 mM Tris(2‐carboxyethyl) phosphine, 0.5% carrier ampholytes 4–6.5, 0.25% carrier ampholytes 3‐10) proved to be very efficient for a large range of different samples and allowed us to obtain two‐dimensional gels of high resolution and quality.