An experimental workflow for enrichment of low abundant proteins from human serum for the discovery of serum biomarkers.

Serum contains proteins that possess important information about diseases and their progression. Unfortunately, these proteins, which carry the information in the serum are in low abundance and are masked by other serum proteins that are in high abundance. Such masking prevents their identification and quantification. Therefore, removal of high abundance proteins is required to enrich, identify, and quantify the low abundance proteins. Immunodepletion methods are often used for this purpose, but there are limitations in their use because of off-target effects and high costs. Here we presented a robust, reproducible and cost-effective experimental workflow to remove immunoglobulins and albumin from serum with high efficiency. The workflow did not suffer from such limitations and enabled identification of 681 low abundance proteins that were otherwise undetectable in the serum. The identified low abundance proteins belonged to 21 different protein classes, namely the immunity-related proteins, modulators of protein-binding activity, and protein-modifying enzymes. They also played roles in various metabolic events, such as integrin signalling, inflammation-mediated signalling, and cadherin signalling. The presented workflow can be adapted to remove abundant proteins from other types of biological material and to provide considerable enrichment for low-abundance proteins.