gene expression

1. PROTEIN MICROARRAY
K.KRUPA SAGAR
M.Pharm 1st year

2. INTRODUCTION
• Protein micro array(or Protein chip) is a high-
throughput method used to track the interactions
& activities of proteins, to determinate their
functions.
• Protein micro array are rapid, automated,
economical and highly sensitive consuming only
small quantities of samples and reagents.

3. HISTORY
• Micro array technology was first developed from
an earlier concept called “Ambient analyte
immunoassay” by Roger Ekins in 1989.
• Later transformed into DNA microarray.
• Due to some limitations in DNA microarray ,
this protein microarray was developed to
overcome those limitations.

4. PRINCIPLE

5. • Protein chip consists of a support surface such
as glass slide, nitrocellulose, bead or
microtitreplate to which array of capture
proteins is bound.
• Probe molecules typically labeled with a
fluorescent dye are added to the array.
• Any reaction between the probe and the
immobilized protein emits a fluorescent
signal and that is read by laser scanner.

6. TYPES OF ARRAY’S:
• Antigen capture
format
• Sandwich format
1.ANALYTICAL
PROTEIN
MICROARRAY
• Biochemical properties
of proteins such as
binding activities =>
• P-P,P-DNA,P-Lipid,
• P-Peptide
2.FUNCTIONAL
PROTEIN
MICROARRAY

7. ANALYTICAL PROTEIN MICROARRAY

8. FUNCTIONAL PROTEIN
MICROARRAY

9. • Functional protein microarray (target protein
array) are constructed by immobilizing large
num of proteins to assay enzymatic activity and
to detect antibodies & their functions.
• They differ from analytical protein microarray
by containing full length functional proteins or
protein domains.

10. • These are used to study the biochemical
activities of entire proteome in a single
experiment.
• The proteins must retain their native or original
structure so that meaningful functional
interactions takes place on the array surface.

11. 3.REVERSE PHASE PROTEIN
MICROARRAY

12. DETECTION SYSTEMS
LABEL - DEPENDENT LABLE- FREE
Several types of labeling
reagents like fluorescent
dyes(Cy3& Cy5),
enzymes(Horseradish peroxides),
radio isotopes (32P, 33P & 14C),
liposome's are used.
Rolling circle amplication (RCA)
Tyramide signal amplication(TSA)
used to detect low abundance
proteins.
Label dependent may effect the
protein activity.
Surface Plasmon Resonance
Spectroscopy (SPRS)
Optical Elipsometry (OE)
Reflectometric Interference
Spectroscopy(RIFS)
Oblique-incidence reflectivity
difference(OIRD)

13. APPLICATIONS
• Mainly used in 5 major areas
• Diagnostics
• Proteomics
• Protein functional analysis
• Antibody characterization &
• Treatment development

14. •Diagnostics:
Detection in antigen antibodies in blood sample
To discover new disease biomarkers; monitoring the
diseased state and responses to therapy.
Ex: Digital bioassay
Proteomics:
Protein expression profiling i.e., which proteins are
expressed in a particular cell.

15. Protein functional analysis:
To identify :-
Protein-protein interactions
Protein- phospholipids interactions
Enzymatic substrates &
Receptor ligands

16. Antibody characterization:
characterization of cross reactivity, specificity and
mapping epitopes.
Treatment development:
 Development of antigen- specificity therapies for
autoimmunity , cancer and allergies.
 Identification of small molecules that could potentially
used to be as new drugs