BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSION: This ELISA method may be useful for rapid screening of anti-AAV-2 neutralizing antibodies in candidates for gene therapy.
BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSION: This ELISA method may be useful for rapid screening of anti-AAV-2 neutralizing antibodies in candidates for gene therapy.
Authors: Kathrina Quinn; Mary R Quirion; Chia-Yun Lo; Julia A Misplon; Suzanne L Epstein; John A Chiorini Journal: Mol Ther Date: 2011-08-09 Impact factor: 11.454
Authors: Matthew R Gardner; Desiree E Mendes; Claudia P Muniz; José M Martinez-Navio; Sebastian P Fuchs; Guangping Gao; Ronald C Desrosiers Journal: Mol Ther Methods Clin Dev Date: 2022-01-07 Impact factor: 6.698
Authors: Joseph Tellez; Kim Van Vliet; Yu-Shan Tseng; Jonathan D Finn; Nick Tschernia; Graça Almeida-Porada; Valder R Arruda; Mavis Agbandje-McKenna; Christopher D Porada Journal: PLoS One Date: 2013-09-24 Impact factor: 3.240