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 function on a large scale. Its main advantage lies in the fact that large numbers of proteins can be tracked in parallel.

1. Romil Jayantilal Mistry
Roll No. 10
M.Sc. B.T. (8th Sem)

2. • Conceptually, protein microarrays are similar to DNA
microarrays.
• They consist of large numbers of proteins individually
immobilized in known positions on the coated surface
of glass slide or silicon chip.
• The proteins arrayed can be antibodies specific for
each protein in an organism, purified recombinant
proteins, or short synthetic peptides.
• There are many ways of attaching a protein to a
support surface.
• The major objective of any coupling system is
maintenance of protein structure and function.
Protein Microarray

3. • Some systems bind proteins to a chemical group that
coat the surface of the support.
• With other protocols, recombinant proteins are
prepared with a short amino acid sequence (tag) at N
or C terminus that bind to a recognition sequence on
the support. In this case, all the protein molecules
are uniformly oriented.
• Instead of spotting proteins on a flat surface, some
microarrays are engineered with tiny depression
(nanowells) that keep each protein moist and prevent
mixing with adjacent proteins.

4. • The purpose of protein microarray analyses is to
detect, on a large scale, the molecules that a protein
interacts with.
• These interacting molecules can be other proteins,
nucleic acid sequences, or low molecular-weight
compounds.
• Protein populations from different samples can be
compared, for example, in control versus treated
samples or in normal versus diseases tissues.

5. • Direct labeling – to label the test samples directly with
a fluorescent dye and then detect the labeled
molecules that bind to the proteins of a microarray
with a laser scanner. Two-dye strategy (e.g. Cy3 or
Cy5) can be used to compare proteins in two different
sample on a single array.
• Sandwich style – the sample molecules are
biotinylated, and after the initial incubation, a
streptavidin-fluorescent-dye conjugate that binds to
biotin to facilitated the detection of sample molecules
is applied.
Protein Microarray - Visualizing

6. Templin et al. 2002 Trend in Biotch. Vol 20

7. Protein array detection method

8. http://www.nature.com/nature/journal/v422/n6928/images/nature01512-f1.2.jpg

9. Protein microarrays
 Many different types
 Antibody arrayed - detect many proteins
 Proteins arrayed - detect interacting proteins
 Proteins arrayed - detect interacting small
molecules , etc.

10. Protein:Protein Interactions

11. Protein activity arrays

12. Small molecule arrays

13. • Cytokine antibody microarrays are used to examine
cytokines in both normal and diseased states, and from a
variety of sources after various treatments.
• A sandwich immunoassay is often used to detect
cytokines that bind to immobilized antibodies.
• After the microarray is treated, biotynylated cytokine
antibodies are added and bind to the corresponding
captured cytokine.
• For visualization, a streptavidin-fluorescent-dye conjugate
attaches to the biotin of the secondary antibody.
• The signals are detected with a laser scanner.