Histo-proteomic profiling of formalin-fixed, paraffin-embedded tissue - PubMed (original) (raw)

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Histo-proteomic profiling of formalin-fixed, paraffin-embedded tissue

Kant M Matsuda et al. Expert Rev Proteomics. 2010 Apr.

Abstract

In the functional proteome era, the proteomic profiling of clinicopathologic-annotated tissues is an essential step for mining and evaluating candidate biomarkers for disease. For many diseases, but especially cancer, the development of predictive biomarkers requires performing assays directly on the diseased tissue. The last decade has seen the explosion of both prognostic and predictive biomarkers in the research setting but few of these biomarkers have entered widespread clinical use. Previously, application of routine proteomic methodologies to clinical formalin-fixed and paraffin-embedded tissue specimens has provided unsatisfactory results. In this paper, we will discuss recent advancements in proteomic profiling technology for clinical applications. These approaches focus on the retention of histomorphologic information as an element of the proteomic analysis.

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Conflict of interest statement

Financial & competing interests disclosure

The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

No writing assistance was utilized in the production of this manuscript.

Figures

Figure 1.

Figure 1.. Components of multiplex tissue immunoblotting.

Diagram showing the transfer ‘package’ utilized for transfer of proteins from a glass slide to membranes of tissue immunoblotting. The package is assembled, then sealed in a Kapak SealPAK® pouch. The package is placed on a heat block and then applied to a multiple-serial heating system (1 h at 55°C, 0.5 h at 65°C and 2 h at 80°C). A weight applied to the top of the transfer assembly unit helps to ensure a tight connection between the layers of material used in the transfer system. The proteins are transferred from the tissue section and are deposited onto the membrane stack. After transfer, the package is disassembled, the membrane stack separated and the proteins on each membrane are individually probed by means of conventional protein blotting.

Figure 2.

Figure 2.. Protein expression in progression from normal to invasive cancer as determined by tissue immunoblotting.

(A) Routine hematoxylin and eosin stain section of esophageal cancer (indicated as an area labeled with ‘T’) along with associated dysplastic (indicated as a triangle area labeled with ‘D’) and normal epithelium (indicated as a rectangle area labeled with ‘N’). (B) CK-4, (C) CK-14, (D) cyclooxygenase-1, (E) annexin 1 and (F) secreted protein acidic and rich in cysteine. Fluorescent scans represented with pseudocolor, where signal intensity is white–red–yellow–green–blue–black from maximum to minimum signal. Modified from [11].

Figure 3.

Figure 3.. Correlation among high-resolution MRI-hisotopathology-multiplex tissue immunoblotting in identical planes.

(A) Ex vivo T2*-MRI of removed human breast tissue at in-plane resolution of 25 × 25 μm. (B) Routine hematoxylin and eosin section. (C) Multiplex tissue immunoblotting for HER2. High-intensity signal is expressed in red color.

Figure 4.

Figure 4.. Schematic of potential application of multiplex tissue immunoblotting in correlation studies with molecular imaging.

Breast specimen is used as an example in this schematic presentation of the work-flow for the correlation study. The arrow indicates co-registration or overlay among radiological imaging (MRI, PET and CT), histopathology section (whole-mount section or regular-size section) and protein expression by MTIB. Initially, in vivo imaging, such as molecular PET imaging for the targeted molecule, is performed, and co-registered with other imaging studies (CT or MRI). Following surgical intervention, ex vivo imaging of the removed tissue is performed and co-registered with in vivo imaging. The targeted lesion, which is seen in in vivo imaging, will be obtained through image-guided sectioning/sampling for histopathologic examination. Once identical planes are obtained, protein expression can be demonstrated using MTIB and readily be correlated with in vivo molecular imaging. MTIB: Multiplex tissue immunoblotting.

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