N‐Glycosylation analysis of formalin fixed paraffin embedded samples by capillary electrophoresis

Abstract

In this study, N-linked glycans from intact, formalin treated and formalin fixed paraffin embedded (FFPE) standard glycoproteins, human serum and mouse tumor tissue samples were investigated in respect to their susceptibility for formaldehyde treatment mediated changes. FFPE samples were first deparaffinized, followed by solubilization in radioimmunoprecipitation assay buffer and treated with PNGase F for N-glycan release. The released glycans were labeled with a charged fluorophore and analyzed by capillary electrophoresis with laser induced fluorescent detection. No significant alterations were found in the N-glycome profile at any of the investigated complexation levels (i.e., glycoprotein, serum and tissue samples) of the study. These results suggest that FFPE samples can be readily used for global N-glycome analysis holding the promise to find novel carbohydrate biomarkers in prospective and retrospective studies. Exoglycosidase based carbohydrate sequencing was also applied to reveal some basic structural information about the N-linked carbohydrates of the mouse tumor tissue samples.

Molecular glycopathology by capillary electrophoresis: Analysis of the N‐glycome of formalin‐fixed paraffin‐embedded mouse tissue samples

“…Our previous work suggested that the FFPE treatment apparently did not affect the glycan moieties of glycoproteins . As a continuation of that work, the objective of this study was to extend the utilization of FFPE samples towards molecular glycopathology for global N‐glycosylation profiling and sequencing with exoglycosidase enzymes.…”

Capillary Electrophoresis: Trends and Recent Advances

“…CGE-LIF was used to identify possible protein N-glycosylation alterations due to FFPE. 163 The N-linked sugars of FFPE treated samples were released using PNGase F digestion. No significant changes were revealed in the N-glycome profile upon FFPE, indicating that global N-glycome analysis on the FFPE samples might be feasible.…”

Biomedical analysis of formalin-fixed, paraffin-embedded tissue samples: The Holy Grail for molecular diagnostics

“…Everest-Dass and Briggs [104] developed a novel method using MALDI MSI on formalin- between the N-glycosylation patterns before and after the formalin fixation and paraffin embedding processes in all sample types examined. Thus, it was suggested that FFPE tissue samples could be readily utilized to discover disease related N-glycosylation changes at the molecular, cellular and tissue levels in prospective and retrospective studies [106].…”