"Identification of protein biomarkers is important because a drug directly acts on proteins," commented Jerry Feitelson, Ph.D., manager of strategic marketing at Beckman Coulter (www.beckmancoulter.com). "Protein biomarkers are generally more predictive compared to genetic or transcriptional biomarkers." They can be used to characterize candidate drugs and are extremely valuable in clinical applications to diagnose disease and monitor therapies.
Protein biomarker research requires analysis of complex biological mixtures that vary in concentration. The biomarker of interest is more often than not present in very low quantities in these samples, relative to the thousands of other proteins present.
"Partitioning and fractionation are two key steps for sample analysis to enrich candidate biomarker proteins,"explained Hans Dewald, product manager for protein enrichment products. "Passage of human sample biofluids, such as plasma, serum, or CSF, through Beckman Coulter's ProteomeLab IgY chemistry partitions 12 of the highly abundant proteins, enriching medium- to low-abundant proteins.
"As opposed to depletion strategies, partitioning allows the captured proteins to be eluted off for additional investigation. We use polyclonal IgY antibodies for the partitioning process because they generate higher avidity capture, have higher specificity, and can be used cross species. This approach can handle up to 250-L sample load, resulting in 1-2 mg of enriched protein of interest per cycle."
Once the samples are partitioned, they can be further enriched through proteome fractionation in two dimensions. The Beckman Coulter ProteomeLab PF 2D is a liquid-fractionation system that allows high mass loads to be fractionated by pH and hydrophobicity. This enables the subsequent identification of proteins at much lower abundance levels, while preserving post-translational modifications, both of which are likely biomarker candidates.
The PF 2D generates liquid fractions of the proteome, as well as pH/hydrophobicity maps, that allow users to visually identify regions of differential protein expression. "Our customers have successfully applied this technology to protein biomarkers, eight orders of magnitude deeper than human serum albumin," said Dr. Feitelson.