Analyzing antibody sequence for recombinant antibody expression_v3

35 Pages·2015·1.63 MB·English

Analyzing antibody sequence for
recombinant antibody expression
Hangxing Yu, Ph.D
Senior Scientist, GenScript
May 20, 2015
Presentation Outline
Antibody basics, structure and function
1
Antibody production in vivo
2
Antibody database and analysis
3
Optimization of antibody production
4
Make Research Easy 2
Antibody basics, structure and function
1
Antibody production in vivo
2
Antibody database and analysis
3
Optimization of antibody production
4
Make Research Easy 3
What is an Antibody?
(cid:1) An antibody (aka Immunoglobulin) is a large Y-shaped protein
produced by B cells.
(cid:1) Used by the immune system to neutralize foreign objects.
Make Research Easy 4
Types of Antibody in vivo
Light Heavy
Name Function
Chain Chain
κ or λ a aaa 1
IgA Found in mucosal areas
κ or λ a aaa 2
Antigen receptors on B
IgD κ or λ δ
cells
Binds to allergen and
IgE κ or λ ε
triggers histamine release
Secreted from B cells with
IgM κ or λ µ
very high avidity
γ1 γ1
Majority of antibody-based
IgG γ2 γ2a
immunity κ or λ
γ3 γ2b
γ4 γ3
Make Research Easy 5
Structural and Functional Units of IgG
(cid:1) "Y"-shaped molecule
(cid:1) Fab region determines its
antigen specificity.
(Fv region and CDR)
(cid:1) Fc region determines the
effects.
(cid:1) Hinge region.
Make Research Easy 6
Antibody-Antigen Interaction
(cid:1) Each antibody binds only to a specific antigen.
(cid:1) Ab-Ag interaction are based on non-covalent binding between Ab
and Ag.
(cid:1) The hinge region allow better binding.
Make Research Easy 7
Antibody-Antigen Interaction
(cid:1) Antibody and antigen interact by spatial
complementarity.
(cid:1) Weak and non-covalent bonds.
electrostatic interactions
hydrogen bonds
Van der Waals forces
hydrophobic interactions
(cid:1) Binding between antibody and antigen is
reversible.
Make Research Easy 8
Complementarity Determining Regions (CDRs)
(cid:1) Ab binding residues are mainly
located in CDRs.
(cid:1) 6 CDRs form the combining site:
L1, L2, L3, H1, H2, H3.
(cid:1) CDR-H3 and L3 have a distinctive
role in antigen recognition.
Make Research Easy 9
Antibody basics, structure and function
1
Antibody production in vivo
2
Antibody database and analysis
3
Optimization of antibody production
4
Make Research Easy 10