One-Step Preservation of Phosphoproteins and Tissue Morphology at Room Temperature for Diagnostic and Research Specimens

BACKGROUND: There is an urgent need to measure phosphorylated cell signaling proteins in cancer tissue for the individualization of molecular targeted kinase inhibitor therapy. However, phosphoproteins fluctuate rapidly following tissue procurement. Snap-freezing preserves phosphoproteins, but is unavailable in most clinics and compromises diagnostic morphology. Formalin fixation preserves tissue histomorphology, but penetrates tissue slowly, and is unsuitable for stabilizing phosphoproteins. We originated and evaluated a novel one-step biomarker and histology preservative (BHP) chemistry that stabilizes signaling protein phosphorylation and retains formalin-like tissue histomorphology with equivalent immunohistochemistry in a single paraffin block. RESULTS: Total protein yield extracted from BHP-fixed, routine paraffin-embedded mouse liver was 100% compared to snap-frozen tissue. The abundance of 14 phosphorylated proteins was found to be stable over extended fixation times in BHP fixed paraffin embedded human colon mucosa. Compared to matched snap-frozen tissue, 8 phosphoproteins were equally preserved in mouse liver, while AMPKβ1 Ser108 was slightly elevated after BHP fixation. More than 25 tissues from mouse, cat and human specimens were evaluated for preservation of histomorphology. Selected tissues were evaluated in a multi-site, independent pathology review. Tissue fixed with BHP showed equivalent preservation of cytoplasmic and membrane cytomorphology, with significantly better nuclear chromatin preservation by BHP compared to formalin. Immunohistochemical staining of 13 non-phosphorylated proteins, including estrogen receptor alpha, progesterone receptor, Ki-67 and Her2, was equal to or stronger in BHP compared to formalin. BHP demonstrated significantly improved immunohistochemical detection of phosphorylated proteins ERK Thr202/Tyr204, GSK3-α/β Ser21/Ser9, p38-MAPK Thr180/Tyr182, eIF4G Ser1108 and Acetyl-CoA Carboxylase Ser79. CONCLUSION: In a single paraffin block BHP preserved the phosphorylation state of several signaling proteins at a level comparable to snap-freezing, while maintaining the full diagnostic immunohistochemical and histomorphologic detail of formalin fixation. This new tissue fixative has the potential to greatly facilitate personalized medicine, biobanking, and phospho-proteomic research

Three Approaches in Group Decision Problems

Majority of approaches in group decision consist in aggregating many individual decisions into one group decision. In this paper a different approach is proposed which is based on the idea of representation of a variety of decisions by a single decision, which could be considered as a most typical. representing the all essential features of given decisions. Such a decision is referred to as a compromise decision. Goodness of representation is defined from three different points of view. All these approaches are equivalent, provided some mild conditions are satisfied

Molecular signatures that can be transferred across different omics platforms

Motivation: Molecular signatures for treatment recommendations are well researched. Still it is challenging to apply them to data generated by different protocols or technical platforms. Results: We analyzed paired data for the same tumors (Burkitt lymphoma, diffuse large B-cell lymphoma) and features that had been generated by different experimental protocols and analytical platforms including the nanoString nCounter and Affymetrix Gene Chip transcriptomics as well as the SWATH and SRM proteomics platforms. A statistical model that assumes independent sample and feature effects accounted for 69-94% of technical variability. We analyzed how variability is propagated through linear signatures possibly affecting predictions and treatment recommendations. Linear signatures with feature weights adding to zero were substantially more robust than unbalanced signatures. They yielded consistent predictions across data from different platforms, both for transcriptomics and proteomics data. Similarly stable were their predictions across data from fresh frozen and matching formalin-fixed paraffin-embedded human tumor tissue

Pressure-Assisted Protein Extraction: A Novel Method for Recovering Proteins from Archival Tissue for Proteomic Analysis

[Image: see text] Formaldehyde-fixed, paraffin-embedded (FFPE) tissue repositories represent a valuable resource for the retrospective study of disease progression and response to therapy. However, the proteomic analysis of FFPE tissues has been hampered by formaldehyde-induced protein modifications, which reduce protein extraction efficiency and may lead to protein misidentification. Here, we demonstrate the use of heat augmented with high hydrostatic pressure (40,000 psi) as a novel method for the recovery of intact proteins from FFPE mouse liver. When FFPE mouse liver was extracted using heat and elevated pressure, there was a 4-fold increase in protein extraction efficiency, a 3-fold increase in the extraction of intact proteins, and up to a 30-fold increase in the number of nonredundant proteins identified by mass spectrometry, compared to matched tissue extracted with heat alone. More importantly, the number of nonredundant proteins identified in the FFPE tissue was nearly identical to that of matched fresh-frozen tissue