Home › Cell Biology › Proteomic Sample Preparation from Formalin Fixed and Paraffin Embedded Tissue
Cell Biology JoVE (Open Access) Citable · DOI

Proteomic Sample Preparation from Formalin Fixed and Paraffin Embedded Tissue

DOI: 10.3791/50589-v
What you'll learn
  • ✓ Prepare microdissected FFPE tissue samples for proteomic analysis
  • ✓ Execute peptide extraction and quantification from archival samples
  • ✓ Fractionate Lys-C and tryptic peptides for downstream proteomics
Protocol

Biopharma Insights Archival formalin fixed and paraffin embedded (FFPE) clinical samples are valuable material for investigation of diseases. Here we demonstrate a sample preparation workflow allowing in-depth proteomic analysis of microdissected FFPE tissue.

Difficulty
advanced
Total time
~4–6 hours per sample (tissue preparation through fractionation)
Biosafety
BSL-1

Steps

1
Prepare tissue slices for microdissection and lysis

Cut FFPE tissue sections and prepare specimens for microdissection. Perform initial tissue lysis to extract protein from archived clinical samples.

▶ 01:46
2
Process samples through protein extraction workflow

Apply chemical and enzymatic treatments to solubilize and denature proteins from FFPE tissue lysates, preparing them for proteomic analysis.

▶ 03:20
3
Quantify extracted peptides by spectroscopy

Measure peptide concentration in processed samples using appropriate quantification assays to standardize subsequent proteomic steps.

▶ 04:51
4
Fractionate Lys-C and tryptic peptides

Separate digested peptide mixtures using liquid chromatography or similar fractionation techniques to reduce complexity for mass spectrometry analysis.

▶ 05:38
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