Journal: ACS Nano

Article Title: Re-Engineered Pseudoviruses for Precise and Robust 3D Mapping of Viral Infection

doi: 10.1021/acsnano.3c07767

Figure Lengend Snippet: Super-resolution imaging of clickVSV. (a) Dual-color d STORM of clickVSVs stained via immunolabeling with α-spike-S2-CF568 (cyan) and click-labeling with Tet-Cy5 (magenta). (b) Particle analysis via LOCAN to study envelope protein density and distribution on the virus surface. (c) Number of S proteins on clickVSVs detected via immunolabeling (gray) and click-labeling (magenta). (d) Particle averaging to resolve the clickVSV shape and intensity. Scale bars: 200 nm (dual-color d STORM) and 100 nm (particle analysis and averaging) (b, d).

Article Snippet: Primary mouse anti-S2-Spike antibody (Thermo Fisher Scientific, MA5-35946) was transferred into 100 mM NaHCO 3 in PBS utilizing Zeba Spin Desalting Columns 40K MWCO (Thermo Fisher Scientific, no. 87766) according to the manufacturer suggested protocol.

Techniques: Imaging, Staining, Immunolabeling, Labeling, Particle Size Analysis, Virus

Journal: ACS Nano

Article Title: Re-Engineered Pseudoviruses for Precise and Robust 3D Mapping of Viral Infection

doi: 10.1021/acsnano.3c07767

Figure Lengend Snippet: ClickVSVs enable visualization of pseudoviruses in different modalities. (a) Live-cell time-lapse imaging of viral infection via 3D lattice-light-sheet (LLS) imaging on HEK293T:ACE2mGreenLantern (grays) cells at different time points after VSVΔG-eGFP-SARS-CoV-2-S Cy5 (magenta) exposure. (b) 3D single particle tracking in two colors shows viral trajectories during infection of HEK293T:ACE2. Here displayed as an XY -maximum intensity projection of all tracks (Tet-ATTO643, magenta; α-spike-S2-CF568, gray). (c) Single trajectory from part b (green dashed box), tracked in both color channels and mapped in a 3D plot showing temporal progression of a particle’s movement on the host cell membrane (Tet-ATTO643, magenta; α-spike-S2-CF568, gray). (d) Track length distribution in two-color tracking experiments with clickVSVs stained with Tet-ATTO643 (magenta, n Tracks = 930) and immunolabeling of α-spike-S2 (gray, n Tracks = 146). (e) PDF of individual diffusion constants of Tet-ATTO643 (magenta), α-spike-S2-CF568 (gray), and fluorescent beads suspended in agarose (light blue). Scale bar, 10 μm (a).

Article Snippet: Primary mouse anti-S2-Spike antibody (Thermo Fisher Scientific, MA5-35946) was transferred into 100 mM NaHCO 3 in PBS utilizing Zeba Spin Desalting Columns 40K MWCO (Thermo Fisher Scientific, no. 87766) according to the manufacturer suggested protocol.

Techniques: Imaging, Infection, Single-particle Tracking, Membrane, Staining, Immunolabeling, Diffusion-based Assay