Development of AAV serotype-specific ELISAs using novel monoclonal antibodies.
Adeno-associated viruses (AAV) have been developed and evaluated as recombinant vectors for gene therapy. More recently, due to the advantages they offer for gene transfer, several AAV serotypes have gained increasing interest. However, monoclonal antibodies for the characterization and quantitation of vectors derived from different serotypes are at present not available. Serotype-specific monoclonal antibodies (mAbs) against the capsids of the serotypes 1/6, 4 and 5 are described. These antibodies, designated as ADK1a and b, ADK4 or ADK5a and b, reacted specifically with the indicated serotype capsids in cell lysates. ADK 1a and b cross-reacted with its highly related AAV6 serotype, but not with the other serotypes tested. The new antibodies recognized exclusively assembled capsids and neither free nor denatured capsid proteins as shown by fractionation experiments. In immunofluorescence experiments, the mAbs stained only distinct intranuclear foci in cells expressing the capsid protein. The development of capture ELISAs for quantitation of AAV1 and 6, AAV4 or AAV5 capsids illustrates that these novel monoclonal antibodies provide valuable tools for characterization of vector stocks.
Kuck
Dirk
D
German Cancer Research Center, Program Infection and Cancer, Heidelberg, Germany.
Kern
Andrea
A
Kleinschmidt
Jürgen A
JA
D000911:Antibodies, Monoclonal / Q000276:immunology*; D002213:Capsid / Q000378:metabolism; D002460:Cell Line; D003429:Cross Reactions; D000229:Dependovirus / Q000145:classification / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005455:Fluorescent Antibody Technique; D006801:Humans; D010957:Plasmids; D012703:Serotyping
D000911:Antibodies, Monoclonal