Neuroantibodies: molecular cloning of a monoclonal antibody against substance P for expression in the central nervous system.
We present a strategy to study functional and/or developmental processes occurring in the nervous system, as well as in other systems, of mice. This strategy is based on the local expression of specific monoclonal antibodies (mAbs) by cells of the nervous system. As an application of this strategy, we report the cloning of the anti-substance P rat mAb NC1/34HL. Functional substance P-binding antibodies were reconstituted from the cloned variable domains by using vectors for expression in myeloma cells. With these and other vectors a general system for the cloning and expression of mAbs under a series of promoters (of the rat VGF8a gene, the neurofilament light-chain gene, and the methallothionein gene) has been created. The activity of these plasmids was confirmed by expressing the recombinant NC1/34HL mAb in GH3 pituitary cells, PC12 pheochromocytoma cells, and COS cells. DNA from the described constructs can be used to target the expression of the NC1/34HL mAb to the central nervous system of transgenic mice. This procedure will allow us to perturb substance P activity in a controlled way in order to dissect its multiple roles.
Piccioli
P
P
Consiglio Nazionale delle Ricerche, Istituto di Neurobiologia, Rome, Italy.
Ruberti
F
F
Biocca
S
S
Di Luzio
A
A
Werge
T M
TM
Bradbury
A
A
Cattaneo
A
A
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics* / Q000276:immunology; D000918:Antibody Specificity; D001483:Base Sequence; D002490:Central Nervous System / Q000502:physiology; D003001:Cloning, Molecular; D015870:Gene Expression; D005803:Genes, Immunoglobulin; D005818:Genetic Engineering; D005822:Genetic Vectors; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D007135:Immunoglobulin Variable Region / Q000235:genetics; D007145:Immunoglobulin kappa-Chains / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D051381:Rats; D013373:Substance P / Q000502:physiology*
D000911:Antibodies, Monoclonal; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D007145:Immunoglobulin kappa-Chains; D013373:Substance P