Applications of Protein Array in Diagnos

tics, Genomic and Proteomics

Microarray technology can simultaneously analyze thousands of
parameters in a single experiment. Micro-point of capture molecules
are fixed into ranks on a solid support and exposed to samples
containing corresponding binding molecules. Complex formation in
each micro-point can be detected by the readout system, which is
based on fluorescence, chemiluminescence, mass spectrometry,
radioactive or electrochemistry. Miniaturization and parallelization
binding assays, whose analysis power can be also enlarged by
microarray gene expression analysis, is sensitive. These systems can
be used to detect the degree of hybridization and immobilized DNA
microarray probes will be exposed to complementary target.
Currently, the development of protein array has demonstrated its
applications in enzyme-substrate, DNA- protein and different types of
protein - protein interactions. In this post, we will discuss the capturemolecule-ligand analysis, analyze its theoretical advantages and
disadvantage and its influence in diagnostics, genomic and
proteomics.

 Theoretically, any kind of ligand-binding assay, which is depending on the pro

duct formation of immobilized capture molecules and the target existing in th
e surrounding solution can be miniaturized and parallelized. This process can
be performed in microarray. The nucleic acid - nucleic acid interaction, which
is also called DNA chips, has been well established, while protein array analysi
s just begins. This can be reflected in recent nucleic acid - protein, protein - pr
otein, ligand - receptor and enzyme - substrate microarray analysis articles.

It is Bulyk and his colleagues who found the application of microarray in DNAprotein interaction. They created the double-stranded oligonucleotide microa
rray. High concentrations of single-stranded oligonucleotide microarray is pro
duced by using Affymetrix (Santa Clara, CA, USA) technology. Single-stranded
oligonucleotide microarray was converted into a double-stranded oligonucleo
tide microarrays under the influence of enzymes extension reaction. Generall
y speaking, DNA- protein interaction analysis is useful for characterizing and i
dentifying DNA-binding proteins, such as, transcription factors.

 There are many kinds of enzyme to analyze enzyme-substrate. In a

conceptual proof experiments, MacBeath and Schreiber fixed three
types of kinase substrate on planar glass surface. Each microarray,
which corresponds to an individual kinase, incubates with radioacti
vely labeled ATP. Each substrate can only be phosphorylated by its s
pecific kinase. In an advanced experiment, Zhu and his colleagues
detected the activity of 119 different protein kinases, which is from
the Saccharomyces cerevisiae, in 17 different substrates. They used
a plate with microwells, in which the substrate interacted. Kinase, w
hich is expressed into GST fusion proteins, is incubated with substr
ate and radiolabeled ATP in microwells. After kinase reaction, the ki
nase and ATP will be washed away, while the array uses phosphatei
mager to analyze the phosphorylated substrate. The new activity of
single kinase can be identified with this method.

 As for receptor - ligand analysis, the small organic molecules produced by the so
lid phase chemistry combination were immobilized in a microarray. The single re
sin beads from combinatorial synthesis are placed in 96-well plates, from which
organic molecules are released chemically. Organic molecules are diluted, disper
sed into small dots, and covalently attached on the slide. These target protein mi
croarrays generated by the so-called small-molecules printing are incubated wit
h the target protein with fluorescently labeled.

In the field of protein - protein interaction analysis, dot-blot filter analysis is use
d for screening the interactions between immobilized protein specific and other
proteins. Some interactions, which happens between radioactively labeled huma
n p52 GST fusion proteins and immobilized capture proteins, have been detecte
d, such as, nuclear proteins, serine - arginine protein fragments isolated from He
La cells. Whats more, this technology has revealed the DNA, RNA, or the interact
ion between low molecular weight ligands with immobilized molecules.

 Recent work of Zhu and his colleages proved the great power of microarray in prot
eomic field. After the purification of 5800 different kinds of recombinant proteins f
rom S. Cerevisiae, the complex protein arrays containing 90 percent of the microbi
al gene were generated. These protein arrays can be used to study the whole- gen
ome-wide protein-protein interactions. Using calmodulin as model protein to prob
e arrays can confirm a number of known interactions and measure a range of nov
el binding proteins. Experiment detecting protein - lipid interaction convincingly ex
plains the possibility of detecting the protein, which can bind with low-molecularweight complexes.

We use microarray technology to screen antigen - antibody interactions. Autoimm

une diseases, such as, system rheumatism, can be diagnosed with 18 different anti
gens, which were immobilized in a microarray. In 1 ml serum in patients, we can d
etect the high accuracy antibody titers. Sandwich immunoassay is also miniaturize
d and parallelled, and may be done in microarray. This has been proved by the diff
erent levels of cytokines in biological samples. This will ultimately lead to powerful
and reliable diagnostic analysis.