High-throughput protein-purification pipelines are becoming part of many proteomics efforts. But how to choose the targets to put through the pipeline? For those who want to focus on targets relevant to a disease-related process, it is important to catalogue where, when and to what extent a protein is expressed. Currently the main method for determining the protein complement of a given cell or tissue uses two-dimensional (2D) gel electrophoresis or liquid chromatography to resolve the proteins in the sample, and mass spectrometry to then identify the individual proteins. A problem with this approach is that it is difficult to identify rare proteins, because cell extracts are dominated by a few very abundant proteins.

Beckman Coulter has developed a number of innovations for protein fractionation to address this problem. “We first selectively remove the most abundant proteins from a sample,” says Jerry Feitelson, marketing manager for the company. The ProteomeLab IgY-12 partitioning kits selectively remove the 12 most abundant proteins — albumins and immunoglobulins — from human/primate serum or plasma. These proteins make up 95% of the total protein expressed by cells. So removing them increases the chances of identifying proteins that are not highly expressed and are probably more interesting to researchers. The technology uses antibodies bound to inert beads, which are then packed in liquid-chromatography columns or, for smaller samples, spin columns. “The advantages of our reagents are the greater capacity, increased specificity and the ability to bind across species,” says Feitelson. The enriched material can then be collected and further fractionated with the Beckman Coulter ProteomeLab PF 2D protein fractionation system. The instrument divides proteins in two dimensions: first, in a 2D gel, samples are separated by isoelectric focusing and size, and then automatically injected into a liquid-chromatography column. The process is fully automated. “One run takes ten hours, so you can do two in one day,” says Feitelson. Detailed protein maps can be constructed for easy comparison between two samples using the Proteome Lab software suite. Interesting proteins can then be further analysed by cutting out the corresponding band, for example, and putting it into a mass spectrometer.

Credit: H. DEWALD/BECKMAN COULTER

Pall Corporation has also introduced the Enchant Life Sciences kits for albumin depletion and immunoglobulin G purification.

L.B.