Figure 1.

AAV panel: Production, quality control, and screening procedure. (a) Workflow of the production and usage of the AAV panel. Fifty AAV capsid variants harboring a CMV-eGFP expression cassette were individually produced and purified over iodixanol gradients. After confirming the quality of all viral batches, vectors were arrayed on 96-well plates and applied for the transduction of various cell types. Following a 1- to 3-day incubation, the percentage of GFP-expressing cells was quantified by high-content confocal imaging and, in some instances, flow cytometry. (b) The production efficiency of each AAV variant was determined by ddPCR and is shown as vg per cm² of culture area. The dotted line indicates the average production efficiency across all batches. AAV genome integrity (expected size: 2,141 bp) was assessed by agarose gel electrophoresis using 1 × 10¹⁰ copies of the extracted scAAV genome per lane. Axis labeling: base pairs (middle panel). Capsid protein identity and purity were assessed by SDS-PAGE and Oriole staining. Axis labeling: kDa (lower panel). (c) AAV panel layout and exemplary high-content imaging output for MIO-M1 cells, 3 days after transduction. AAV, adeno-associated virus; ddPCR, droplet digital polymerase chain reaction; sc, self-complementary; SDS-PAGE, sodium dodecyl sulfate–polyacrylamide gel electrophoresis.