Fig. 3: Functional evaluation of AAV vectors in the ex situ human liver perfused with non-neutralizing human plasma.

a Graphical representation of the study. b Concentration of total (top panel) and de-multiplexed AAV vector genomes in the perfusate (bottom panel). Concentration of individual vectors is expressed as the estimated percentage of the initial concentration as studied with NGS. Vectors are ranked from most to least abundant (top to bottom). c Total vector genomes per haploid cell found in biopsies from both liver grafts extracted at the indicated perfusion times. d Percentage of NGS reads mapped to each barcoded AAV capsid variant. The transgene DNA, indicating vector uptake, was extracted from biopsies taken from both grafts at the indicated time points. e Similar analysis performed on transgenes recovered from RNA, which indicate functional transduction. Percentages are normalized to the pre-injection mix. f Immunofluorescence analysis of the net eGFP signal from collective AAV transduction in the left graft. g Similar immunofluorescence image for the right graft. The vector-encoded eGFP was also counterstained with an anti-eGFP antibody (red). Blue: DAPI (nuclei). Scale = 100 μm. h Immunofluorescence analysis of the bile ducts and hepatocytes of the left graft. Yellow: Cytokeratin 7 (bile ducts); red: albumin; blue: DAPI (nuclei). Scale: 50 μm. i Immunofluorescence analysis of liver zonation. Red: Histidine ammonia-lyase (HAL); blue: DAPI (nuclei). Scale: 500 μm. j Immunofluorescence analysis of liver integrity. Red: albumin; Purple: Hepatocyte nuclear factor 4 alpha (HNF4α); blue: DAPI (non-hepatocyte nuclei). Scale: 20 μm. k Immunofluorescence analysis of AAV Receptor (AAVR) in the human liver. Orange: AAVR; Blue: DAPI (nuclei). Scale: 100 μm.