Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2023 Jan 2;41(7):932–943. doi: 10.1038/s41587-022-01567-w

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

PMC Copyright notice

Fig. 4 — a, Schematic representation of the BiTE construct. Clone 13 anti-Trp2 scFv was used for one binding module, and the scFv of the anti-CD3 Ab 2C11 was used for the T cell-binding module. b, scFv13 BiTE showed cytotoxic activity against Trp2 peptide-pulsed EL4 cells. CFSE and PI double-positive cells were gated and the experiment was triplicated. Dead cell percentage shown as mean ± s.d. (n = 3 biological replicates). c, scFv13 BiTE showed cytotoxic activity against B16F10 cells. B16F10 cells were pretreated (+I) or not (–I) with IFN-γ and pulsed (+P) or not (–P) with Trp2 peptide. Treated B16F10 cells were then incubated with mouse T cell blasts at varying concentrations of BiTE or 2C11 scFv. CFSE and PI double-positive cells were gated and the experiment was triplicated. Dead cell percentage shown as mean ± s.d. (n = 3 biological replicates). d, Nonspecific BITE activity derives from 2C11 Ab to CD3, not from Trp2 TCRm. Trp2-negative EL4 cells were pulsed (+P) or not (–P) with Trp2 peptide and then incubated with mouse T cell blasts at different concentrations of BiTE (BiTE) or 2C11 (2C11) scFv. CFSE and PI double-positive cells were gated and the experiment was triplicated. Dead cell percentage are shown as mean ± s.d. (n = 3 biological replicates). e, Schematic representation of the CAR-T experiment. f, scFv13-CAR-T specifically kills B16F10 cells. B16F10 cells were either incubated alone (B16 only); pretreated with IFN-γ (B16+IFNg); incubated with CAR-T (B16 + T cell); pretreated with IFN-γ and incubated with CAR-T (B16 + T cell + IFNg); or pretreated with INF-γ, pulsed with Trp2 peptide and incubated with CAR-T (B16 + T cell + IFNg + peptide). The E:T ratio varied between 1:1, 3:1 and 10:1. B16F10 cell death was monitored by DAPI staining. Representative dead cell percentage shown as mean ± s.d. (n = 3 biological replicates). g, scFv13 is specific for Trp2-expressing cells. CAR-T or CD19 CAR-T cells were incubated with mouse B cells at E:T ratios of 1:1 and 3:1. Live B cell numbers were directly counted in the DAPI-negative gate and are shown as mean ± s.d. (n = 3 biological replicates).

Source data