Two expression vectors for the phage-displayed chicken monoclonal antibody.
We previously reported the development of chicken monoclonal antibodies (mAb) against mammalian-conserved molecules by cell fusion and phage display using the mouse mAb expression vector pPDS. However, chicken hybridomas produce relatively small amounts of antibody when compared with mouse hybridomas, and application of the pPDS may be limited in two-antibody assays with a mouse mAb because it contains mouse Ckappa as a detection tag. To circumvent the above problems, two expression vectors were established and used to produce a functional recombinant chicken mAb. These vectors, which were designed to accommodate a single chain fragment of the variable region (scFv) of the antibody, contained a chicken Clambda and FLAG with or without 6 x histidine sequences in the 3' terminus of the scFv to serve as detection and purification tags. In this study, a prion protein (PrP)-specific chicken mAb (HUC2-13) was expressed as phage-displayed and soluble scFv mAb forms by using these vectors. The scFv mAbs expressed by these vectors exhibited the same antigen-binding specificity to PrP as that of the original HUC2-13, could be purified with ease, and used in combination with a mouse mAb. These results indicate that the methods described herein offer an alternative to chicken mAb production from hybridomas and immunized chicken splenocytes, and may contribute to the use of chicken mAb reagents in numerous fields.
Nakamura
Naoto
N
Laboratory of Immunobiology, Department of Molecular and Applied Biosciences, Graduate School of Biosphere Sciences, Hiroshima University, 1-4-4 Kagamiyama, Higashi, Hiroshima 739-8528, Japan.
Shimokawa
Mariko
M
Miyamoto
Kazuyoshi
K
Hojyo
Shintaro
S
Horiuchi
Hiroyuki
H
Furusawa
Shuichi
S
Matsuda
Haruo
H
D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics* / Q000302:isolation & purification; D000918:Antibody Specificity; D001483:Base Sequence; D002645:Chickens / Q000235:genetics* / Q000276:immunology*; D018076:DNA, Complementary / Q000235:genetics; D005822:Genetic Vectors; D006825:Hybridomas / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000302:isolation & purification; D051379:Mice; D019151:Peptide Library; D011328:Prions / Q000276:immunology; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification
D000911:Antibodies, Monoclonal; D018076:DNA, Complementary; D007135:Immunoglobulin Variable Region; D019151:Peptide Library; D011328:Prions; D011994:Recombinant Proteins