Journal: Molecular therapy. Methods & clinical development

Article Title: Unveiling the secrets of adeno-associated virus: novel high-throughput approaches for the quantification of multiple serotypes.

doi: 10.1016/j.omtm.2023.101118

Figure Lengend Snippet: Figure 1. Genomic titers of different dPCR and qPCR methods VG titers of AAV2, AAV5, and AAV8 reference standard materials (AMSBIO Europe, Alkmaar, the Netherlands) were determined by cdPCR, ndPCR, ddPCR, mapdPCR, and qPCR in a duplex assay targeting both the SV40 and ITR region on the AAV genome. The VG titers for either the SV40 or the ITR region are displayed for AAV2 (A and D), AAV5 (B and E), and AAV8 (C and F). Error bars represent SDs of independent triplicate measurements. Statistical analysis was performed using one-factor ANOVA with a sig- nificance level of a = 0.05. Statistically significant difference is denoted by an asterisk, and “ns” indicates no significant difference. vg, viral genome; SV40, simian virus 40, ITR, inverted terminal repeats.

Article Snippet: The active working time er 2023 VG titers of AAV2, AAV5, and AAV8 reference standard materials (AMSBIO Europe, Alkmaar, the Netherlands) were determined by cdPCR, ndPCR, ddPCR, mapdPCR, and qPCR in a duplex assay targeting both the SV40 and ITR region on the AAV genome.

Techniques: Virus

Journal: Molecular therapy. Methods & clinical development

Article Title: Unveiling the secrets of adeno-associated virus: novel high-throughput approaches for the quantification of multiple serotypes.

doi: 10.1016/j.omtm.2023.101118

Figure Lengend Snippet: Figure 3. Sample preparation for capsid lysis Serially diluted AAV2, AAV5, and AAV8 samples were lysed either by a Proteinase K digest or by heat incubation. Re- sulting VG titers are presented exemplifying the SV40 ndPCR on the QIAcuity One (Qiagen). Error bars represent SDs of independent triplicate measurements. VG titers were analyzed using an unpaired t test with a significance level of a = 0.05, which is indicated by an asterisk. vg, viral genome; SV40, simian virus 40; +, with; -, without.

Article Snippet: The active working time er 2023 VG titers of AAV2, AAV5, and AAV8 reference standard materials (AMSBIO Europe, Alkmaar, the Netherlands) were determined by cdPCR, ndPCR, ddPCR, mapdPCR, and qPCR in a duplex assay targeting both the SV40 and ITR region on the AAV genome.

Techniques: Sample Prep, Lysis, Incubation, Virus

Journal: Molecular therapy. Methods & clinical development

Article Title: Unveiling the secrets of adeno-associated virus: novel high-throughput approaches for the quantification of multiple serotypes.

doi: 10.1016/j.omtm.2023.101118

Figure Lengend Snippet: Figure 4. Capsid titer determination and handling times of capsid ELISA and BLI A correlation between capsid titer and absorbance, as well as between capsid titer and binding rate, for ELISA (A) and BLI (B), is shown for AAV8. Furthermore, the determined capsid titers for AAV2, AAV5, and AAV8 samples, obtained from two distinct upstream processes, within the linear range by both methods are presented (C). In addition, the active as well as the passive working time for both techniques for the processing of 16 samples is shown (D). The active working time involves manual handling steps, such as pipetting, whereas the passive working time refers to waiting times, such as incubation or measurement times. Preparation of 16 samples and their transfer to the respective 96-well plates was identical for both methods and took approximately 1 h. Therefore, this step was excluded from the active working time analysis. Error bars represent SDs of replicate measurements (ELISA, n = 2; BLI, n = 3). Capsid titers were analyzed using an unpaired t test with a significance level of a = 0.05. Significant differences are represented by an asterisk, and “ns” denotes no significant difference. Batch A and batch B, respective upstream process; t, time.

Article Snippet: The active working time er 2023 VG titers of AAV2, AAV5, and AAV8 reference standard materials (AMSBIO Europe, Alkmaar, the Netherlands) were determined by cdPCR, ndPCR, ddPCR, mapdPCR, and qPCR in a duplex assay targeting both the SV40 and ITR region on the AAV genome.

Techniques: Enzyme-linked Immunosorbent Assay, Binding Assay, Incubation