Fig 1.
After sample collection, both blood and liver are processed. Left lateral liver lobe is cut and digested on a GentleMACS Dissociator followed by centrifugation separation of hepatocytes and intra-hepatic cells. Lymphocytes are purified using a Percoll gradient to remove debris and remaining hepatocytes. Hepatocytes are pelleted for debris removal. After cell counting, cells are ready for all downstream applications. The blood samples are collected from the saphenous vein, after a filtering step, blood is incubated with magnetic beads for red blood cell depletion. After washing, cells are counted and ready for downstream applications. Image created with BioRender.
Fig 2.
Flow cytometry to identify liver resident cell population composition after enzymatic digestion (right) compared to mechanical disruption (left).
(a) Graphical representation of cell populations theoretically present in mouse liver (Image created with BioRender) (b) Dot plot for living (APC-Cy7) CD45+ (BUV395) cells shows an extra population of CD45 negative cells (indicated with an arrow). (c) Dot plot showing the Kupffer cell population (indicated with arrow) which are identified as CD11b+ (BUV496) and F4/80+ (PE-Dazzle594) double positive cells of living CD45+ population. (d) Pie chart showing cell populations within the living CD45+ cells (n = 5).
Fig 3.
Single-cell RNA sequencing data for comparison of mechanical with enzymatic digestion of mouse liver.
(a) UMAP projection of the cell population annotations across the two liver dissociation methods. (b) Bar plot with cell population proportions across the two liver dissociation methods. (c) Bar plot with cell population proportions across the different samples.
Table 1.
Cell numbers distribution of the different cell populations per liver isolation protocol.
The aggregated sum per cell type for each protocol is shown An overview of the cell type annotations can be found in S5 Fig.
Fig 4.
Primary mouse hepatocytes can be used for antiviral compound testing using confocal imaging and HBsAg CLIA.
(a) Culture of primary mouse hepatocytes from AAV-HBV infected mice can be maintained for at least 14 days and (b) can be used for ex vivo compound screening of antiviral compounds which show an inhibition of HBsAg in the supernatant of the hepatocyte culture in three independent experiments and (c) induced speckling of HBc visualized by confocal imaging in which Hoechst is visualized in blue, HBsAg is colored in orange (CF568) and core protein is stained in green (AlexaFluor488).
Fig 5.
Single-cell sequencing data from comparison of protocols for purification of whole blood.
(a) UMAP projection of cell types across the different whole blood purification methods (b) Bar plot with cell population proportions across the different whole blood purification methods. (c) Dot plot visualization of scaled expression of the contamination of the different red blood cell markers per protocol. The size of the dots represents the percentage of cells expressing the marker across all cells from the condition. An overview of the cell type annotations can be found in S9 Fig.
Table 2.
Cell number distribution of the difference cell populations per blood isolation protocol.
The aggregated sum per cell type for each protocol is shown, an overview of the cell type annotations can be found in S9 Fig.
Fig 6.
Overview of possible downstream applications after isolation of mouse liver and blood cells.