Analysis of cloned Fvs from a phage display library indicates that DNA immunization can mimic antibody response generated by cell immunizations.
BACKGROUND
Generation and cloning of antibodies against cell surface antigens can be simplified by combining DNA immunization which enables generation of antibodies against a protein in its natural configuration without the need for any protein purification step and antibody phage display which due to its immense screening power and physical coupling between the phenotype and genotype of antibodies simplifies the cloning of antibody genes.


OBJECTIVE
Since DNA immunization is expected to elicit antibodies against a protein in its natural configuration, we wanted to see if it can mimic the antibody response generated by cell immunization.


METHODS
A phage display library made from splenic mRNA of a mouse immunized with mesothelin cDNA was panned on mesothelin-positive cells. The single-chain Fvs (scFvs) selected were then analyzed.


RESULTS
We obtained several anti-mesothelin scFvs. One of these Fvs is almost identical to the Fv of a monoclonal antibody that was previously obtained from a hybridoma in which the mice were immunized with a mesothelin-positive ovarian cancer cell line. Another Fv was found to be specific for mesothelin present on human cells.


CONCLUSIONS
Our results indicate that an antibody phage display library made from spleens of DNA-immunized mice is a rapid and efficient alternative to cell immunization for obtaining antibodies against different epitopes of a membrane antigen that is very difficult to purify in a native form.
Chowdhury
P S
PS
National Institutes of Health, National Cancer Institute, Laboratory of Molecular Biology, Bethesda, MD 20892-4255, USA.
Pastan
I
I
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D058851:GPI-Linked Proteins; D006801:Humans; D007114:Immunization / Q000379:methods; D007128:Immunoglobulin Fragments / Q000276:immunology*; D007135:Immunoglobulin Variable Region / Q000276:immunology; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000276:immunology*; D000090204:Mesothelin; D051379:Mice; D008807:Mice, Inbred BALB C; D018716:Molecular Mimicry; D008969:Molecular Sequence Data; D019151:Peptide Library; D011993:Recombinant Fusion Proteins / Q000276:immunology; D014407:Tumor Cells, Cultured; D019444:Vaccines, DNA / Q000276:immunology*
D058851:GPI-Linked Proteins; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D008562:Membrane Glycoproteins; C000715528:Msln protein, mouse; D019151:Peptide Library; D011993:Recombinant Fusion Proteins; D019444:Vaccines, DNA; D000090204:Mesothelin