Workflow for Combined Proteomics and Glycomics Profiling from Histological Tissues

Extracellular
matrixes comprise glycoproteins, glycosaminoglycans
and proteoglycans that order the environment through which cells receive
signals and communicate. Proteomic and glycomic molecular signatures
from tissue surfaces can add diagnostic power to the immunohistochemistry
workflows. Acquired in a spatially resolved manner, such proteomic
and glycomic information can help characterize disease processes and
be easily applied in a clinical setting. Our aim toward obtaining
integrated omics datasets was to develop the first workflow applicable
for simultaneous analysis of glycosaminoglycans, N-glycans and proteins/peptides from tissue surface areas as small
as 1.5 mm in diameter. Targeting small areas is especially important
in the case of glycans, as their distribution can be very heterogeneous
between different tissue regions. We first established reliable and
reproducible digestion protocols for the individual compound classes
by applying standards on the tissue using microwave irradiation to
achieve reduced digestion times. Next, we developed a multienzyme
workflow suitable for analysis of the different compound classes.
Applicability of the workflow was demonstrated on serial mouse brain
and liver sections, both fresh frozen and formalin-fixed. The glycomics
data from the 1.5 mm diameter tissue surface area was consistent with
data published on bulk mouse liver and brain tissues, which demonstrates
the power of the workflow in obtaining combined molecular signatures
from very small tissue regions.