Discovery of modulators of B cell receptor response using a calcium mobilization assay.

Abstract

The B cell receptor (BCR) induces a sustained increase in cytoplasmic free Ca2+ upon activation. This ensures the progression of calcium-dependent signaling processes such as protein phosphorylation, gene transcription, cell cycle progression, or apoptosis. In an effort to identify and characterize potential immunomodulatory agents we have used freshly isolated and Ramos B cells to establish a FLIPR-based method for measuring effects of various agents on BCR-mediated Ca2+ signaling. Here we report on a Ca2+ assay that allows sensitive and robust detection of inhibitory actions of both mAbs as well as inhibitors of downstream tyrosine kinase targets. The observed effects on Ca2+ signaling have been corroborated with FACS analyses of develomental and proliferative functions. Our method can be applied to the systematic evaluation of potential drugs which may modify B cell development, activation or proliferation through effects on Ca2+ signaling.