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. 2021 Jun 2;13(6):1055. doi: 10.3390/v13061055

Figure 1.

Figure 1

Construction and validation of RBD:ACE2 NanoBiT biosensor. (A) Schematic of SRAE2-BS constructs. SP, signal peptide. (B) Demonstration of SRAE2-BS working mechanism. (C) Western blot analysis of protein expression of SRAE2 components. Plasmids were transfected into HEK293T cells in 12-well plate. 10 µg of protein lysates were subjected to SDS-PAGE, followed by western blot analysis using anti-His first antibody. b-actin was used as internal protein loading control. Molecular weight in kilodalton (kDa) is shown on the right of the blot. (D) Luciferase analysis of different combinations of biosensor components. Plasmids were transfected into HEK293T cells in 96-well plate, followed by luciferase analysis using cell lysates from proteins extracted from transfected cells (upper panel) or secreted protein from cell culture medium (lower panel). RLU, relative luciferase unit. (E) Luciferase analysis of SRAE2-BS activity in vitro. Equal amounts (0–10 µg) of Sm-ACE2-His and RBD-Lg-His protein lysates were mixed and incubated together for 30 min, followed by luciferase analysis.