Journal: bioRxiv

Article Title: NewroBus for the brain: humanized TfR1-targeting nanobodies with high BBB permeability and cargo transport capacity

doi: 10.1101/2025.04.20.649139

Figure Lengend Snippet: A ) Schematic representation of the 16 humanized and optimized TNFI–TfR1b heterodimers. B ) Flow cytometry analysis in HEK293 cells stably expressing human TfR1, comparing the binding of heterodimers to that of their parental humanized TfR1b-Nbs. C ) Dose-response apoptosis inhibition curves in WEHI-13VAR cells treated with human TNFα and 2-fold serial dilutions of TNFI–TfR1b heterodimers (100,000 to 12.21 pM). Apoptosis was measured using a fluorogenic caspase-3/7 activation assay. The IC₅₀ values of the heterodimers were comparable to those of the parental humanized nanobodies TNFI-α (IC₅₀ = 48.84 pM, range: 42.11–56.56) and TNFI-β (IC₅₀ = 64.31 pM, range: 57.17–72.31).

Article Snippet: The same procedure was used with the following modifications: plates were coated with 0.2 µg/mL biotinylated human TNFα (Acro Biosystems, TNA-H82E3-25ug) instead of TfR1.

Techniques: Flow Cytometry, Stable Transfection, Expressing, Binding Assay, Inhibition, Activation Assay