Proteomics-Based Analysis and Diagnosis of Formalin-Fixed Paraffin-Embedded Amyloidosis Samples.

Abstract

Amyloidosis is a group of rare pathologies characterized by abnormal folding and deposition of susceptible proteins in tissues and organs. Diagnosis of amyloidosis often relies on immunohistochemistry of formalin-fixed paraffin-embedded (FFPE) patient samples; however, dependency on antibodies for protein staining is one of the major pitfalls of this approach, especially for the detection of rare amyloidosis types. In recent years, mass spectrometry-based proteomics has emerged as a promising alternative for adequate detection and amyloid typing, despite the fact that preparing FFPE samples for proteomics remains a challenging task. Major hurdles are removal of formalin-induced protein cross-links and water-insoluble paraffin prior to mass spectrometry analysis. With the recent development of the suspension trapping protocol, enabling the use of high concentrations of SDS, these obstacles can be overcome. In this chapter, we describe the implementation of suspension trapping for FFPE sample processing and its application to analyze human amyloidosis samples, comparing a standard procedure with probe sonication with a more advanced workflow based on ultrasonication.