A convenient enzyme-linked immunosorbent assay for rapid screening of anti-adeno-associated virus neutralizing antibodies

• Published in: Annals of clinical biochemistry Vol. 46; no. Pt 6; p. 508
• Main Authors: Ito, Tetsuo, Yamamoto, Shigekazu, Hayashi, Tsukasa, Kodera, Mika, Mizukami, Hiroaki, Ozawa, Keiya, Muramatsu, Shin-ichi
• Format: Journal Article
• Language: English
• Published: England 01.11.2009
• Subjects: Antibodies, Neutralizing - chemistry; beta-Galactosidase - metabolism; Cell Line; Chemistry, Clinical - methods; Dependovirus - genetics; Enzyme-Linked Immunosorbent Assay - methods; Gene Transfer Techniques; Genetic Therapy - methods; Genetic Vectors; Horseradish Peroxidase - genetics; Humans; Immunoglobulin G - genetics; Neutralization Tests; Reference Values
• ISSN: 1758-1001
• DOI: 10.1258/acb.2009.009077

Recombinant adeno-associated virus vectors based on serotype 2 (AAV-2) have become leading vehicles for gene therapy. Most humans in the general population have anti-AAV-2 antibodies as a result of naturally acquired infections. Pre-existing immunity to AAV-2 might affect the functional and safety consequences of AAV-2 vector-mediated gene transfer in clinical applications. An enzyme-linked immunosorbent assay (ELISA) method was developed using microwell plates coated with intact particles of recombinant AAV-2 vectors, and horseradish peroxidase-conjugated anti-human immunoglobulin G (HRP-IgG). Neutralizing antibody titres were analysed by assessing the ability of serum antibody to inhibit transduction into HEK293 cells of AAV vectors that express beta-galactosidase. Anti-AAV-2 antibodies were detected by ELISA in two of 20 healthy subjects. The positivity criterion (optical density >0.5) in ELISA corresponded to the cut-off value (320-fold dilution of serum) in the AAV-2 neutralization assay. Influences of interfering substances were not observed. This ELISA method may be useful for rapid screening of anti-AAV-2 neutralizing antibodies in candidates for gene therapy.