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4 Paik et al.
2. Sample Specimens and Processing Techniques Used for Clinical
Proteomics
2.1. General Considerations
Because clinical proteomics rely heavily on the patient specimens, three
important factors need to be considered before the selection and preparation of
clinical specimens: (1) selection of the correct clinical samples according to the
type of research, (2) isolation of the appropriate component from the clinical
samples, and (3) establishment of optimal experimental conditions for each
sample (5,6,7,8). For the selection of correct clinical samples, the relationship
between clinical samples and the specific disease should also be considered.
For example, although cancer tissue represents a specific cancer, several types
of body fluids from patients may also have a relationship to the cancer. If
the selected clinical samples specifically represent the disease, the next step
is to evaluate what components are related to the specific disease. That is,
tumor cells in cancerous tissues are surrounded by many types of stromal cells,
inflammatory cells, and connective tissues that are directly related to changes
in protein expression in the cancer. If the purpose of proteomic analysis is
to identify characteristic changes of specific proteins in tumor cells, then the
precise identification of tumor cell percentage that can be increased by tissue
microdissection would appear to be necessary (5,6,7). As sample specimen
conditions directly impact the results of biomarker discovery, well-defined
clinical specimens should be used since the discovery of disease biomarkers is
much easier when the samples have clear anatomical and pathophysiological
definitions. Because clinical specimens are heterogeneous, sophisticated pathological
discrimination is required for the isolation of specific diseased tissue or
body fluids. Without the expertise of a pathologist at the earliest stage, it may
be difficult to isolate a specifically defined specimen for clinical proteomics.
Generally, clinical samples contain variable factors and components originating
from the microenvironment of specific tissues. For instance, liver tissues usually
contain a large amount of blood in the sinusoid and this amount is increased
in tissues with dilated sinusoids (9). Lung tissues usually contain deposited
exogenous materials and this amount is increased in heavy smokers (10). Note
that the amount of blood present in isolated tissues may directly influence the
relative proportion of proteins found in clinical specimens. Deposited materials
and the other chemicals such as stain dye and fixatives used in the microdissection
may also influence the experimental conditions (11). In the analysis of
clinical samples, suitable buffer conditions, minimal lysis time, and high-yield
protein precipitation are highly recommended. To avoid substantial variations
between experiments using clinical specimens, a large set of specimens are
also necessary because, unlike cultured cell lines, clinical specimens have high