Physical titer of FVIII vectors as determined by reverse transcriptase assay and quantitative PCR
| Virus . | Average reverse transcriptase, ng/μL . | Estimated titer, TU/mL . | Titer, vector genomes/mL . |
|---|---|---|---|
| BDD FVIII | 10.9 | 3.71 × 109 | 1.14 × 108 |
| FVIII Fugu B | 46.5 | 1.58 × 1010 | 1.58 × 109 |
| FVIII N6 | 30.7 | 1.04 × 1010 | 1.07 × 109 |
| SQ FVIII | 68.3 | 2.32 × 1010 | 2.91 × 109 |
| SQ FVIII Fugu B | 44.8 | 1.52 × 1010 | 1.18 × 109 |
| SQ FVIII N6 | 78.0 | 2.65 × 1010 | 2.0 × 109 |
| SQ FVIII (co) | 69.6 | 2.37 × 1010 | 4.45 × 109 |
| SQ FVIII Fugu B (co) | 71.8 | 2.40 × 1010 | 2.65 × 109 |
| SQ FVIII N6 (co) | 87.9 | 2.99 × 1010 | 3.39 × 109 |
| Virus . | Average reverse transcriptase, ng/μL . | Estimated titer, TU/mL . | Titer, vector genomes/mL . |
|---|---|---|---|
| BDD FVIII | 10.9 | 3.71 × 109 | 1.14 × 108 |
| FVIII Fugu B | 46.5 | 1.58 × 1010 | 1.58 × 109 |
| FVIII N6 | 30.7 | 1.04 × 1010 | 1.07 × 109 |
| SQ FVIII | 68.3 | 2.32 × 1010 | 2.91 × 109 |
| SQ FVIII Fugu B | 44.8 | 1.52 × 1010 | 1.18 × 109 |
| SQ FVIII N6 | 78.0 | 2.65 × 1010 | 2.0 × 109 |
| SQ FVIII (co) | 69.6 | 2.37 × 1010 | 4.45 × 109 |
| SQ FVIII Fugu B (co) | 71.8 | 2.40 × 1010 | 2.65 × 109 |
| SQ FVIII N6 (co) | 87.9 | 2.99 × 1010 | 3.39 × 109 |
Quantification of RT protein concentration in viral stocks, measured by performing an RT colorimetric assay, quantified in nanograms per microliter and estimated titer calculated from this. Data are mean values; n = 3. Quantification of titer was determined using quantitative PCR. A total of 1 × 105 293T cells were transduced with a serial dilution of vector; after 72 hours, genomic DNA was extracted from cells and quantitative PCR carried out for both WPRE and the human housekeeping gene β-actin. Data are mean values; n = 5.