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Protein Micro Array

A protein microarray provides a multiplex approach to identify protein-protein interactions and other molecular interactions by affixing different proteins or DNA binding sequences to specific locations on a glass slide to form a microscopic array. The most common type is an antibody microarray where antibodies are spotted onto the array to detect proteins from cell lysate solutions. Protein microarrays are measurement devices used in biomedical applications to determine the presence and amount of proteins in biological samples like blood.

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56 views3 pages

Protein Micro Array

A protein microarray provides a multiplex approach to identify protein-protein interactions and other molecular interactions by affixing different proteins or DNA binding sequences to specific locations on a glass slide to form a microscopic array. The most common type is an antibody microarray where antibodies are spotted onto the array to detect proteins from cell lysate solutions. Protein microarrays are measurement devices used in biomedical applications to determine the presence and amount of proteins in biological samples like blood.

Uploaded by

Binil Manuel
Copyright
© Attribution Non-Commercial (BY-NC)
We take content rights seriously. If you suspect this is your content, claim it here.
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Download as DOCX, PDF, TXT or read online on Scribd
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Protein microarray

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A protein microarray, sometimes referred to as a protein binding microarray, provides a


multiplex approach to identify protein–protein interactions, to identify the substrates of protein
kinases, to identify transcription factor protein-activation, or to identify the targets of
biologically active small molecules. The array is a piece of glass on which different molecules of
protein or specific DNA binding sequences (as capture probes for the proteins) have been affixed
at separate locations in an ordered manner thus forming a microscopic array. The most common
protein microarray is the antibody microarray, where antibodies are spotted onto the protein chip
and are used as capture molecules to detect proteins from cell lysate solutions.

Related microarray technologies also include DNA microarrays, cellular microarrays, antibody
microarrays, tissue microarrays and chemical compound microarrays.

Contents
[hide]

 1 Applications
 2 Types of chips
 3 Production of protein arrays
o 3.1 Artifacts to avoid
 4 Types of capture molecules
 5 Detection methods
 6 See also
 7 Software/source code
 8 References
 9 External links

[edit] Applications
Protein microarrays (also biochip, proteinchip) are measurement devices used in biomedical
applications to determine the presence and/or amount (referred to as relative quantitation) of
proteins in biological samples, e.g. blood. They have the potential to be an important tool for
proteomics research. Usually a multitude of different capture agents, most frequently monoclonal
antibodies, are deposited on a chip surface (glass or silicon) in a miniature array. This format is
often also referred to as a microarray (a more general term for chip based biological
measurement devices).
[edit] Types of chips
There are several types of protein chips, the most common being glass slide chips and nano-well
arrays.

[edit] Production of protein arrays


The production process depends on the type of protein chip. protein–protein array: The proteins
can be externally synthesised, purified and attached to the array. Alternatively they can be
synthesised in-situ and directly attached to the array. The proteins can be synthesised through
biosynthesis, cell-free DNA expression or chemical synthesis. In-situ synthesis is possible with
the latter two. With cell-free DNA expression, proteins are attached to the support right after
their production. Peptides chemically procured by solid phase peptide synthesis are already
attached to the support. Selective deprotection is carried out through lithographic methods or by
the so-called SPOT-synthesis.

DNA-Protein array: Double-stranded DNA (the exact binding sequence of the protein)is
attached/ spotted on the array.

[edit] Artifacts to avoid

 1) To avoid variability in results, use a very efficient lysis buffer and maintain consistent
sample processing conditions;
 2) Many antibodies don't work well as capture reagents, even if they do work well in
western blotting and other denaturing conditions. Some antibodies often bind poorly to
intact proteins in a cell extract;
 3) Different proteins like different solution conditions, so if you do not see binding it
doesn't mean that there is no binding between the two partners in physiological
conditions;
 4) Adjust the solute conditions to avoid non-specific association: change salt
concentration, pH, add 1% alignate;
 5) on the array's surface the conjugated protein should be in the right conformation (i.e.,
folded, NOT denatured), anchored by the same amino acid (in the same orientation), and
be kept away from the surface by a linker to avoid steric hindrance.

[edit] Types of capture molecules


Capture molecules used are most commonly antibodies; however, antigens are used in
applications where antibodies are detected in serum. More recently there has been a push
towards other types of capture molecules which are more similar in their nature such as peptides
or aptamers. Antibodies have several problems including the fact that there are no antibodies for
most proteins and also problems with specificity in some commercial antibody preparations.
Nevertheless, antibodies still represent the most well-characterized and effective protein capture
agent for microarrays. Recently, nucleic acids, receptors, enzymes, and proteins have been
spotted onto chips and used as capture molecules. This allows a vast variety of experiments to be
conducted on protein–protein interactions, and all other protein binding substrates.

[edit] Detection methods


Although protein microarrays may use similar detection methods as DNA Microarrays, a
problem is that protein concentrations in a biological sample may be many orders of magnitude
different from that for mRNAs. Therefore, protein chip detection methods must have a much
larger range of detection.

The preferred method of detection currently is fluorescence detection. The fluorescent detection
method is compatible with standard microarray scanners, the spots on the resulting image can be
quantified by commonly used microarray quantification software packages. However, some
minor alterations to the analysis software may be needed. Other common detection methods
include colorimetric technqiues based on silver-precipitation, chemiluminescent and label free
Surface Plasmon Resonance.

[edit] See also

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