In vivo expression of a GST-fusion protein mediates the rapid generation of affinity matured monoclonal antibodies using DNA-based immunizations.
Previously we demonstrated the rapid generation of affinity matured monoclonal antibody (MAb) producing cell lines following gene gun delivery of DNA using a mammalian expression vector (pAlpha/hFc), which enables the expression of human Fc-chimera proteins in vivo. Here we compare the pAlpha/hFc vector to modified vectors that replace human IgG(1) with either a Glutathione-S-Transferase (GST) fusion protein or a mouse IgG(2c) (mFc) fusion protein. We report that in vivo expression of a GST-chimera results in the rapid generation of affinity matured MAbs, comparable with antibodies raised using the pAlpha/hFc vector, that were reactive with annexin V. The mFc vector failed to induce early antigen-specific B-cell responses suitable for MAb development.
MV CEED Biology, GlaxoSmithKline, Research Triangle Park, NC 27709, USA. kkilpatrick@tripathimaging.com
Katherine E
KE
Kilpatrick
Sandy
S
Kerner
Eric P
EP
Dixon
Jeff T
JT
Hutchins
Janet H
JH
Parham
J Patrick
JP
Condreay
Gregory
G
Pahel
D000818:Animals; D017304:Annexin A5 / Q000276:immunology; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D000915:Antibody Affinity; D001483:Base Sequence; D019470:Biolistics; D004274:DNA, Recombinant / Q000235:genetics; D015870:Gene Expression; D005822:Genetic Vectors; D005982:Glutathione Transferase / Q000235:genetics / Q000276:immunology; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D007114:Immunization; D007074:Immunoglobulin G / Q000235:genetics / Q000276:immunology; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology*
D017304:Annexin A5; D000911:Antibodies, Monoclonal; D004274:DNA, Recombinant; D007074:Immunoglobulin G; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase