Thermal Denaturation of Fresh Frozen Tissue Enhances Mass Spectrometry Detection of Peptides

Thermal denaturation
(TD), known as antigen retrieval, heats tissue
samples in a buffered solution to expose protein epitopes. Thermal
denaturation of formalin-fixed paraffin-embedded samples enhances
on-tissue tryptic digestion, increasing peptide detection using matrix-assisted
laser desorption ionization imaging mass spectrometry (MALDI IMS).
We investigated the tissue-dependent effects of TD on peptide MALDI
IMS and liquid chromatography-tandem mass spectrometry signal in unfixed,
frozen human colon, ovary, and pancreas tissue. In a triplicate experiment
using time-of-flight, orbitrap, and Fourier-transform ion cyclotron
resonance mass spectrometry platforms, we found that TD had a tissue-dependent
effect on peptide signal, resulting in a (22.5%) improvement in peptide
detection from the colon, a (73.3%) improvement in ovary tissue, and
a (96.6%) improvement in pancreas tissue. Biochemical analysis of
identified peptides shows that TD facilitates identification of hydrophobic
peptides.