Identification of Notch ligand useage by splenic B cells using Dll1 and Dll4 Notch ligand blockade.

Marginal zone (MZ) B cells leverage Notch2 signals to attain rapid differentiation responses and positional cues with in the spenic environment.To determine the likely molecular source of Notch2 signals we sought to block the Notch ligands delta-like ligand 1 (Dll1) and delta-like ligand 4 (Dll4) in wildtype mice using in-vivo treatment with monoclonal blocking antibodies. Splenic populations of MZ B cells and follicular B cells were sort purified after 12, 24 and 48 hours of blockade and analyzed by RNA-seq. Overall design: C57BL/6 (10-12 weeks of age) were treated with anti-Dll1, anti-Dll4 blocking antibodies or isotype control antibodies for 12-48 hours prior to sacrifice. Marginal zone B cells (CD19+, CD93-, CD1d+, CD23-) and follicular B cells (CD19+, CD93-, CD1d-, CD23+) were purified by FACS sorting and RNA-sequencing library generation.