Differential screening of phage-ab libraries by oligonucleotide microarray technology.
A novel and efficient tagArray technology was developed that allows rapid identification of antibodies which bind to receptors with a specific expression profile, in the absence of biological information. This method is based on the cloning of a specific, short nucleotide sequence (tag) in the phagemid coding for each phage-displayed antibody fragment (phage-Ab) present in a library. In order to set up and validate the method we identified about 10,000 different phage-Abs binding to receptors expressed in their native form on the cell surface (10 k Membranome collection) and tagged each individual phage-Ab. The frequency of each phage-Ab in a given population can at this point be inferred by measuring the frequency of its associated tag sequence through standard DNA hybridization methods. Using tiny amounts of biological samples we identified phage-Abs binding to receptors preferentially expressed on primary tumor cells rather than on cells obtained from matched normal tissues. These antibodies inhibited cell proliferation in vitro and tumor development in vivo, thus representing therapeutic lead candidates.
Monaci
Paolo
P
Biotechnology Department, Istituto di Ricerca di Biologia Molecolare P. Angeletti, Pomezia, Rome, Italy. paolo_monaci@merck.com
Luzzago
Alessandra
A
Santini
Claudia
C
De Pra
Alessandra
A
Arcuri
Mirko
M
Magistri
Francesca
F
Bellini
Alessandro
A
Ansuini
Helenia
H
Ambrosio
Maria
M
Ammendola
Virginia
V
Bigotti
Maria Giulia
MG
Cirillo
Agostino
A
Nuzzo
Maurizio
M
Nasti
Annamaria Assunta
AA
Neuner
Philippe
P
Orsatti
Laura
L
Pezzanera
Monica
M
Sbardellati
Andrea
A
Silvestre
Giuseppe
G
Uva
Paolo
P
Viti
Valentina
V
Barbato
Gaetano
G
Colloca
Stefano
S
Demartis
Anna
A
De Rinaldis
Emanuele
E
Giampaoli
Saverio
S
Lahm
Armin
A
Palombo
Fabio
F
Talamo
Fabio
F
Vitelli
Alessandra
A
Nicosia
Alfredo
A
Cortese
Riccardo
R
D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics* / Q000378:metabolism / Q000493:pharmacokinetics; D001435:Bacteriophages / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay; D007074:Immunoglobulin G / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D020411:Oligonucleotide Array Sequence Analysis; D020349:Surface Plasmon Resonance
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D008565:Membrane Proteins