Fibroblastic reticular cells from secondary lymphoid organs provide Notch signals to promote effector CD8 ⁺ T cell differentiation during infection. Control (Ccl19-Cre⁻-Dll1/4^fl/fl; referred to as Dll1/4^fl/fl) or mice with a specific loss of Notch ligands DLL1 and DLL4 in FRCs (Ccl19-Cre⁺-Dll1/4^fl/fl; referred to as Dll1/4^Δ/Δ) were infected with Lm-OVA. As a comparison, Notch1/2 KO (E8I-Cre⁺Notch1/2^fl/fl) mice were included. The CD8⁺ T cell response was assessed 7 days later in the spleen by flow cytometry. (A) Representative plots (left) and compilation (right) of the OVA-specific polyclonal CD8⁺ T cell response detected by tetramer staining. (B) Representative plots (left) and compilation (right) of the proportion of SLECs, MPECs, and early effector cells (EECs) among Tet-OVA⁺ CD8⁺ T cells. (C) Binding of the 1B11 antibody to detect the core-2 O-glycosylated form of CD43 was used as a Notch signaling indicator on Tet-OVA⁺ cells from Dll1/4^fl/fl and Dll1/4^Δ/Δ mice following Lm-OVA infection. (D) Representative plots (left) and compilation (right) of the proportion of CX3CR1^hiCXCR3^lo SLECs from Dll1/4^fl/fl and Dll1/4^Δ/Δ mice following Lm-OVA infection. (E) WT mice were treated with an isotype control or blocking antibodies against DLL1, DLL4, or both on days 0 and 2 after Lm-OVA infection. The proportion of SLECs was measured among Tet-OVA⁺ CD8⁺ T cells. Data are from two independent experiments with a total of n = 6–8 mice per group (A–E). Error bars display means ± SEM. Ordinary one-way ANOVA with Tukey’s multiple comparison was used for multiple group comparison. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.