The document discusses methods for isolating antibodies from recombinant antibody libraries using various selection platforms that mimic in vivo processes. It outlines different classification methods for selection, including solid-phase, solution-sorting, cell-based, and in vitro-based screening, and describes how elution conditions can enhance selection. Key steps in antibody generation through these methods involve generating diversity, coupling phenotype to genotype, and confirming binding through screening.

1. Antibodies can be isolated
from recombinant antibody
libraries in lab. Using one of
these platforms for selection
that in essence mimics in vivo
process.
Platforms for antibody display and selection1
Antigen presenting strategies3
To carry out the selections
from display libraries, many
different methods and experi-
mental approaches have been
developed to separate clones
that bind from those that do
not. According to the proper-
ties of the phases that hold
the specific antigens, these
selection methods can be
classified into solid-phase,
solution-sorting, cell-based
and in vitro-based screening.
Elution conditions can also be
used to drive the selection
towards the desired popula-
tion, for example, via
trypsin-digestion of a proteo-
lytically sensitive phage, via
mild disulfide-bridge reduc-
tion to release phage or both
antigen and phage, or via
competitive elution with a
ligand binding to the antigen
and displacing the relevant
phage antibody.
Phage Display Library Construction
Immune Antibody Library
Scaffold Library
Constrained Peptide Library
cDNA Library
Genome Library
ANTIBODY LIBRARY SCREENING
Creative Biolabs
Phage Display
Library Screening
Services
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Phage display
Protein-mRNA
link via:
Display on:
-ribosome display
-mRNA display
-Yeast
-Bacteria
-Mammalian cells
-Retroviruses
-.....
Protein-DNA display
Growth selection via:
Microbead via
compartmentalization
-Yeast two-hybrid
-Protein fragment
complementation
Immobilized Ag
Biotinylated Ag
Cells displaying
antigen/internalizing Ag
Eluting strategies4
Mild reduction
(release phage only)
SS
Trypsin digestion
Mild reduction
(release phage +Ag)
SS
Competitive elution
Alternating selections
on Ag+/– cells
Subcellular fractions
Phage Display Library Screening
Solid-Phase Screening
Solution-Sorting Screening
Cell-Based Screening
Screening
Screening
Solid Phase Based
Solution Sorting Based
Cell Based
Tissue with Ag
Ex Vivo Based
Most of the platforms for
antibody selection and
screening share four key steps
with the procedure for
antibody generation in the in
vivo immune system: First,
antibody diversity is generat-
ed from synthetic V genes or
cloned from B cells. Next,
antibody phenotype is
coupled to its genotype via a
phenotype-genotype link
packaged in a host (shown
here schematically for phage
display). As a result, each host
particle expresses a unique
antibody on its surface. The
repertoire of antibodies
displayed on these host parti-
cles is subjected to the
process is repeated and even-
tually antibodies binding to
antigen are confirmed by
screening.
Magnetic beads
coated Ab
Steps in antibody selection and screening2
Coupling
of geno to
phenotype
Selective
pressure on
phenotype
Screening
Antibody
gene pool
Displayed library
Selected antibody lead
Synthetic
DNA
Cloning
of genetic
diversity
B cells
Selection cycle
+
Amplification
Mutagenesis
and selection
cycle