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

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.