Kinetic measurement of AAV vector transduction efficiencies to optimize assay duration

In this experiment, Celigo was used to image and quantify AAV-GFP transduction efficiencies in 96-well plates for 6 different serotypes at high, mid, and low MOIs over time.

1. Target cells are seeded in each well of a 96-well plate and incubated overnight
2. Cells are transduced with six types of AAV-GFP vectors at high, medium, and low MOIs and incubated for overnight transduction
3. The Celigo plate imager imaged and analyzed in brightfield and green fluorescence to measure the transduction efficiencies at three MOIs for different serotypes of AAV-GFP vectors
4. The Celigo analyzed time points at approximately 20, 40, and 60 hours
5. The software automatically identifies and segments cells to provide accurate counts of both total cells and GFP+ cells

Determine changes in transduction efficiencies and gene expression levels with time-course measurements of GFP population and fluorescence intensities

• MOI and time-dependent increases in GFP+ cells were observed for the six different AAV serotypes (Figure 1)
• AAV-C demonstrated the highest efficiency at 20 hours for all three MOIs
• AAV-A and AAV-C had similar percentages of GFP+ percentages at 40 hours
• AAV-A outperformed AAV-C at the final time point
• AAV-B showed higher gene expression over time in comparison to AAV-D (Figure 2)

Figure 1. A time-course plot of six serotypes of AAV vectors at low MOI.

Figure 2. A time-course plot of GFP fluorescent intensities for AAV-B and AAV-D, where AAV-B showed higher gene expression.

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