Viral envelope protein gp64 transgenic mouse facilitates the generation of monoclonal antibodies against exogenous membrane proteins displayed on baculovirus.
We have been investigating the functional display of multipass membrane protein such as transporter or G-protein coupled receptor on the budded baculovirus (BV). We tested the use of a viral envelope protein gp64 transgenic mouse for the direct immunization of these membrane proteins displayed on BVs. The gp64 transgenic mice showed only a weak response to virus compared to wild type BALB/c mice. Immunizing gp64 transgenic mice with the BV expressing peptide transporter PepT1, we obtained 47 monoclonal antibodies (mAbs). These mAbs were specific to the PepT1 on the pancreatic cancer cells AsPC-1 by fluorocytometric analysis and exhibited antibody-dependent cellular cytotoxicity or complement-dependent cytotoxicity to AsPC-1. We also generated 7 mAbs by immunizing gp64 transgenic mice on a CCR2-deficient background with the BV expressing chemokine receptor CCR2 together with partially purified CCR2. These mAbs possessed specific binding to CCR2 in CHO cells on fluorocytometric analysis, and exhibited neutralizing activities for ligand-dependent inhibition of cyclic AMP production. This method provides a powerful tool for the generation of therapeutic/diagnostic mAbs against membrane proteins.
Saitoh
Ryoichi
R
Fuji Gotemba Research Laboratories, Chugai Pharmaceutical Co. Ltd., 1-135 Komakado, Gotemba, Shizuoka, 412-8513, Japan.
Ohtomo
Toshihiko
T
Yamada
Yoshiki
Y
Kamada
Nobuo
N
Nezu
Jun-Ichi
J
Kimura
Naoki
N
Funahashi
Shin-Ichi
S
Furugaki
Koh
K
Yoshino
Takeshi
T
Kawase
Yosuke
Y
Kato
Atsuhiko
A
Ueda
Otoya
O
Jishage
Kou-Ichi
K
Suzuki
Masami
M
Fukuda
Rie
R
Arai
Masahiro
M
Iwanari
Hiroko
H
Takahashi
Kazuaki
K
Sakihama
Toshiko
T
Ohizumi
Iwao
I
Kodama
Tatsuhiko
T
Tsuchiya
Masayuki
M
Hamakubo
Takao
T
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D016367:Baculoviridae / Q000235:genetics* / Q000378:metabolism; D016466:CHO Cells; D015815:Cell Adhesion Molecules / Q000235:genetics*; D045744:Cell Line, Tumor; D006224:Cricetinae; D003412:Cricetulus; D007114:Immunization; D008562:Membrane Glycoproteins / Q000235:genetics*; D008565:Membrane Proteins / Q000235:genetics / Q000276:immunology*; D051379:Mice; D008822:Mice, Transgenic; D019151:Peptide Library; D000074011:Peptide Transporter 1; D054390:Receptors, CCR2; D019707:Receptors, Chemokine / Q000276:immunology; D027981:Symporters / Q000276:immunology; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000276:immunology; D014764:Viral Proteins / Q000235:genetics*
D000911:Antibodies, Monoclonal; C515335:Ccr2 protein, mouse; D015815:Cell Adhesion Molecules; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D019151:Peptide Library; D000074011:Peptide Transporter 1; D054390:Receptors, CCR2; D019707:Receptors, Chemokine; D027981:Symporters; D014759:Viral Envelope Proteins; D014764:Viral Proteins