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Updated: Aug 8, 2026
Proteomic Sample Preparation from Formalin Fixed and Paraffin Embedded Tissue
Published on: September 3, 2013
Release and analysis of polypeptides and glycopolypeptides from formalin-fixed, paraffin wax-embedded tissue
S A Brooks1, M V Dwek, A J Leathem
1School of Biological and Molecular Sciences, Oxford Brookes University, Headington, Oxford, UK.
Insights
Formalin-fixed tissues can now be biochemically analyzed. Cyanogen bromide cleavage solubilizes proteins from archival samples, enabling new research correlations.
Area of Science:
- Biochemistry
- Histology
- Proteomics
Background:
- Archival tissues are formalin-fixed and paraffin-embedded, preserving morphology but cross-linking proteins.
- Formalin fixation makes proteins insoluble, preventing standard biochemical analysis of histochemically identified components.
- This limits correlating tissue biochemistry with long-term clinical follow-up.
Purpose of the Study:
- To develop a method for solubilizing proteins from formalin-fixed, paraffin-embedded tissues.
- To enable biochemical analysis of archival tissue specimens.
- To facilitate correlation of tissue biochemistry with long-term clinical data.
Main Methods:
- Applied cyanogen bromide (CNBr) cleavage to formalin-fixed, paraffin-embedded rat liver tissue.
- CNBr specifically cleaves proteins at methionine residues.
- Analyzed released transferrin polypeptides and glycopolypeptides, including oligosaccharide structures.
Main Results:
- Successfully solubilized transferrin polypeptides and glycopolypeptides from archival tissue.
- Confirmed that sialylated oligosaccharide structures remained intact after CNBr cleavage.
- Demonstrated the feasibility of biochemical analysis on formalin-fixed samples.
Conclusions:
- Cyanogen bromide cleavage is an effective method for solubilizing proteins from formalin-fixed, paraffin-embedded tissues.
- This technique preserves oligosaccharide structures, allowing for detailed biochemical analysis.
- The approach offers broad applicability for research correlating tissue biochemistry with clinical outcomes.
Abstract:
Archival tissue specimens are commonly stored as formalin-fixed, paraffin wax-embedded blocks. Formalin fixation facilitates excellent morphological preservation, and the immunoreactivity of many antigens is preserved, but formalin-induced chemical cross-linking of proteins renders them insoluble and inaccessible to standard biochemical extraction and analytical methods. Thus, biochemical analysis of tissue components identified by histochemistry, with the advantage of long-term clinical follow-up, is precluded. We have applied cyanogen bromide cleavage, a technique used routinely for fragmenting proteins for sequencing experiments, to solubilize transferrin polypeptides and glycopolypeptides from formalin-fixed, paraffin wax-embedded rat liver. Cyanogen bromide cleaves protein specifically at methionine residues, yielding a predictable array of polypeptide fragments. Subsequent oligosaccharide analysis of the transferrin glycopolypeptides by anion exchange chromatography confirmed that, in addition to successful release of polypeptide chains, sialylated oligosaccharide structures remained intact after cyanogen bromide cleavage. This approach may have wide applicability to a range of research interests in which correlation of tissue biochemistry with long-term follow-up is advantageous.
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