Applications of Protein Array in Diagnostics, Genomic and Proteomics

1
Applications of Protein Array in Diagnostics,
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
  capture-molecule-ligand analysis, analyze its
  theoretical advantages and disadvantage and its
  influence in diagnostics, genomic and proteomics.

2

• Theoretically, any kind of ligand-binding assay,
  which is depending on the product formation of
  immobilized capture molecules and the target
  existing in the 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 analysis just begins. This can be
  reflected in recent nucleic acid - protein,
  protein - protein, ligand - receptor and enzyme -
  substrate microarray analysis articles.
• It is Bulyk and his colleagues who found the
  application of microarray in DNA- protein
  interaction. They created the double-stranded
  oligonucleotide microarray. High concentrations
  of single-stranded oligonucleotide microarray is
  produced by using Affymetrix (Santa Clara, CA,
  USA) technology. Single-stranded oligonucleotide
  microarray was converted into a double-stranded
  oligonucleotide microarrays under the influence
  of enzymes extension reaction. Generally
  speaking, DNA- protein interaction analysis is
  useful for characterizing and identifying
  DNA-binding proteins, such as, transcription
  factors.

3

• 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 radioactively
  labeled ATP. Each substrate can only be
  phosphorylated by its specific 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, which is expressed into GST
  fusion proteins, is incubated with substrate and
  radiolabeled ATP in microwells. After kinase
  reaction, the kinase and ATP will be washed away,
  while the array uses phosphateimager to analyze
  the phosphorylated substrate. The new activity of
  single kinase can be identified with this method.
  

4

• As for receptor - ligand analysis, the small
  organic molecules produced by the solid phase
  chemistry combination were immobilized in a
  microarray. The single resin beads from
  combinatorial synthesis are placed in 96-well
  plates, from which organic molecules are released
  chemically. Organic molecules are diluted,
  dispersed into small dots, and covalently
  attached on the slide. These target protein
  microarrays generated by the so-called
  small-molecules printing are incubated with the
  target protein with fluorescently labeled.
• In the field of protein - protein interaction
  analysis, dot-blot filter analysis is used for
  screening the interactions between immobilized
  protein specific and other proteins. Some
  interactions, which happens between radioactively
  labeled human p52 GST fusion proteins and
  immobilized capture proteins, have been detected,
  such as, nuclear proteins, serine - arginine
  protein fragments isolated from HeLa cells.
  Whats more, this technology has revealed the
  DNA, RNA, or the interaction between low
  molecular weight ligands with immobilized
  molecules.

5

• Recent work of Zhu and his colleages proved the
  great power of microarray in proteomic field.
  After the purification of 5800 different kinds of
  recombinant proteins from S. Cerevisiae, the
  complex protein arrays containing 90 percent of
  the microbial gene were generated. These protein
  arrays can be used to study the whole-
  genome-wide protein-protein interactions. Using
  calmodulin as model protein to probe arrays can
  confirm a number of known interactions and
  measure a range of novel binding proteins.
  Experiment detecting protein - lipid interaction
  convincingly explains the possibility of
  detecting the protein, which can bind with
  low-molecular-weight complexes.
• We use microarray technology to screen antigen -
  antibody interactions. Autoimmune diseases, such
  as, system rheumatism, can be diagnosed with 18
  different antigens, which were immobilized in a
  microarray. In 1 ml serum in patients, we can
  detect the high accuracy antibody titers.
  Sandwich immunoassay is also miniaturized and
  parallelled, and may be done in microarray. This
  has been proved by the different levels of
  cytokines in biological samples. This will
  ultimately lead to powerful and reliable
  diagnostic analysis.