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Protein Microarray

A protein microarray is a high-throughput method to study protein interactions and function on a large scale. It consists of an array of capture proteins bound to a solid surface, such as a glass slide. Probe molecules are added and any reactions emit fluorescent signals that are scanned. There are three main types of protein microarrays: analytical microarrays use antibodies to detect proteins in samples; functional microarrays contain full-length proteins to study biochemical activities; and reverse phase microarrays apply cell lysates to study disease-related protein alterations.

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578 views5 pages

Protein Microarray

A protein microarray is a high-throughput method to study protein interactions and function on a large scale. It consists of an array of capture proteins bound to a solid surface, such as a glass slide. Probe molecules are added and any reactions emit fluorescent signals that are scanned. There are three main types of protein microarrays: analytical microarrays use antibodies to detect proteins in samples; functional microarrays contain full-length proteins to study biochemical activities; and reverse phase microarrays apply cell lysates to study disease-related protein alterations.

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PROTEIN MICROARRAY

 A protein microarray (or protein chip) is a high-throughput method used to track the
interactions and activities of proteins, and to determine their function, and determining functionon
a large scale.
 Its main advantage lies in the fact that large numbers of proteins can be tracked in parallel.
 The chip consists of a support surface such as a glass slide, nitrocellulose membrane,
bead,or micro-titre plate, to which an array of capture proteins is bound.
 Probe molecules, typically labelled with a fluorescent dye, are added to the array. Any
reaction between the probe and the immobilised protein emits a fluorescent signal that is read by
a laserscanner.
 Protein microarrays are rapid, automated, economical, and highly sensitive, consuming
smallquantities of samples and reagents.
 The concept and methodology of protein microarrays was first introduced and illustrated
in antibody microarrays (also referred to as antibody matrix) in 1983 in a scientific publication
and a series of patents.
 The high-throughput technology behind the protein microarray was relatively easy to
develop since it is based on the technology developed for DNA microarrays, which have become
the mostwidely used microarrays.
Applications :
There are five major areas where protein arrays are being applied:
 Diagnostics
 Proteomics
 Protein functional analysis
 Antibody characterization
 Treatment development.

DIFFERENT TYPES OF PROTEIN MICROARRAY


There are three types of protein microarrays that are currently used to study the biochemical
activities of proteins.
 Analytical protein microarrays
 Functional protein microarrays
 Reverse phase protein microarrays

These types of microarrays can be used to monitor differential expression profiles and for clinical
diagnostics. Examples include profiling responses to environmental stress and healthy versus disease
tissues

Analytical Microarray:
Analytical microarrays (or antibody microarrays) have antibodies arrays on solid surface and are
used to detect proteins in biological samples. Often a second is used to detect a protein that is
captured by the antibody attached to the solid phase, in a principle similar to that of sandwich
immunoassay, in which the first antibody is spotted on the array and then a captured antigen on the
chip is detected with a second antibody that recognises a different part of antigen.

Analytical microarrays are typically used to profile a complex mixture of proteins in order to
measure binding affinities, specificities, and protein expression levels of the proteins in the mixture.
In this technique, a library of antibodies, aptamers, or affibodies is arrayed on a glass microscope
slide. The array is then probed with a protein solution. Antibody microarrays are the most common
analytical microarray
In order to increase affinity and specificity, analytical micro-arrays usually employ a signal
amplification system and sandwich assay format, in which the first antibody is spotted on the array
and then a captured antigen on the chip is detected with a second antibody that recognizes a
different part of the antigen.

Functional Microarray –
It is also known as target protein array.

Functional protein microarrays differ from analytical arrays in that functional protein arrays are
composed of arrays containing full-length functional proteins or protein domains. These protein chips
are used to study the biochemical activities of an entire proteome in a single experiment.

Functional protein microarrays have recently been applied to many aspects of discovery-based
biology, including protein-protein, protein-lipid, protein-DNA, protein-drug, and protein-peptide
interactions. Although we have attempted to describe all the major applications of functional protein
microarrays, it is impossible to cover all the instances in which they have been used.

Different types of biochemical assays were carried out on chips, including assays of

 protein-protein
 protein-lipid
 protein-DNA
 protein-drug
 (H) protein-small molecule
 Protein-antibody interactions.

The chips can also be used to monitor immune responses in patients and posttranslational
modifications of proteins, such as phosphorylation.

These assays achieved high signal-to-noise ratios and were very informative for elucidating the
function of previously uncharacterized genes.
Reverse Microarray –
A third type of protein microarray, related to analytical microarrays, is known as a reverse phase
protein microarray (RPA). In RPA, cells are isolated from various tissues of interest and are lysed. The
lysate is arrayed onto a nitrocellulose slide using a contact pin microarray. The slides are then probed
with antibodies against the target protein of interest, and the antibodies are typically detected with
chemiluminescent, fluorescent, or colorimetric assays. Reference peptides are printed on the slides
to allow for protein quantification of the sample lysates.

It involve complex samples, such as tissue lysates. Cells are isolated from various tissues of interest
and lysed. The lysate is arranged onto the microarray & probed with antibodies against the target
protein of interest. These antibodies are typically detected with chemiluminescent, fluorescent or
colorimetric assays.

RPAs allow for the determination of the presence of altered proteins or other agents that may be
the result of disease. Specifically, post translational modifications, which are typically altered as a
result of disease, can be detected using RPAs.

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