Biology Reference
In-Depth Information
albumin, which is a major contributor to osmotic plasma pressure,
and assists in the transport of fatty acids and steroid hormones (
3
).
Immunoglobulins make up 8-26% of the plasma protein and play
a role in the transport of ions, hormones, and lipids through the
circulation system. Approximately 4% is fi brinogen, which can be
converted into insoluble fi brin and is essential for the clotting of
blood. Regulatory proteins, which make up less than 1% of plasma
protein, include cytokines, enzymes, proenzymes, and hormones.
Current research regarding plasma protein is centered on perform-
ing proteomic analysis of serum/plasma samples to identify disease
biomarkers. Gel-based proteomic approaches rely on reducing the
complexity of whole plasma by depleting high-abundance proteins
with affi nity chromatography (
4
) and/or by using premade IPG
strips within a narrow pH range.
Hepatocellular carcinoma (HCC) is a common cancer and
accounts for nearly 40% of all cancers and approximately 90% of
primary liver cancers in Southeast Asia (
5
). HCC usually develops
in cirrhotic livers that are infected with chronic hepatitis B virus
(HBV), hepatitis C virus (HCV), or coinfected with human immu-
nodefi ciency virus (HIV) and HBV or HCV (
6
). Although HCC
has been the subject of considerable research interest, the associ-
ated prognosis and death rates have remained nearly constant,
which has been attributed to ineffi cient diagnosis. Current tech-
niques for diagnosing HCC involve screening for the presence of
one or more biomarkers including alpha-fetoprotein (AFP), des-
gamma-carboxyprothrombin (DCP), glypican-3 (GPC3), alpha-
L
-
fucosidase (AFU), and transforming growth factor (TGF)-beta1
(
7, 8
). Although these biomarkers have proven useful for detecting
HCC, they generally suffer from limited sensitivity and/or speci-
fi city (
9
). Thus, the development of a new class of biomarkers for
the diagnosis of HCC is an urgent research priority (
10-12
).
In our laboratory, we have previously used various proteomic
techniques, such as two-dimensional electrophoresis (2DE), 2-D
liquid chromatography (LC) coupled to the ProteomeLab Protein
Fractionation System (PF2D), and isotope labeling, to identify dif-
ferences in protein expression between clinical plasma and liver tis-
sue samples (
13, 14
). These proteomic studies suggest that the
characterization of proteins with posttranslational modifi cations
(PTMs) and selection of the optimal proteomic methods are the
key factors that drive the discovery of novel biomarkers (
15-17
).
Although 2D PAGE is the most powerful gel-based method to
separate and visualize proteins, the recognized problems with this
approach are inconsistent gel-to-gel reproducibility and limited
dynamic range due to low sensitivity. An improved method is two-
dimensional fluorescence difference gel electrophoresis (2D
DIGE), in which samples are labeled individually with fl uorescent
cyanine dye (Cy2, Cy3, and Cy5) and then pooled before separa-
tion and scanning in a single gel. This approach overcomes the
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