Proteomics of hydrophobic samples: Fast, robust and low-cost workflows for clinical approaches.

ABSTRACT

In a comparative study, we investigated the influence of nine sample preparation workflows and seven different lysis buffers for qualitative and quantitative analysis of the human adipose tissue proteome. Adipose tissue is not just a fat depot but also an endocrine organ, which cross-talks with other tissue types and organs throughout the body, like liver, muscle, pancreas, and brain. Its secreted molecules have an influence on the nervous, immune, and vascular system, thus adipose tissue plays an important role in the regulation of whole-body homeostasis. Proteomic analysis of adipose tissue is challenging due to the extremely high lipid content and a variety of different cell types included. We investigated the influence of different detergents to the lysis buffer and compared commonly used methods like protein precipitation and filter-aided sample preparation (FASP) with workflows involving acid labile or precipitable surfactants. The results indicate that a sodium deoxycholate (SDC) based workflow had the highest efficiency and reproducibility for quantitative proteomic analysis. In total 2564 proteins from the adipose tissue of a single person were identified.