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Fig. 4. Determination of a reference proteome. As not any desired amount of cells can be
collected by microdissection, a reference proteome was established for subsequent pro-
tein spot identifi cation. 2D DIGE analysis of a mixture of grade 2 and grade 3 tumor tissue
harvested from 25-
μ
m-thick sections (
d
) showed a spot matching rate of >90% to gels
with separated protein from microdissected bronchial epithelium (
a
) and squamous cell
carcinoma tissue (
b
). The matching rate of spot pattern produced from epithelium cells
from esophagus (which can be sampled much more easily) (
c
) was about 80%, and there-
fore esophagus was not considered for subsequent experiments.
9. After scanning, analyze the images using the DIA module of
DeCyder (see Fig.
4
) and calculate the matching rate between
microdissected cells and carcinoma tissue protein spots.
10. Optimize the labeling conditions for the tissue lysate showing
the highest matching rate with the microdissected cells by a
same/same experiment (see Subheading
3.3
).
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