B cell receptor (BCR)-mediated signaling pathways are important for B cell proliferation and differentiation. Abnormal B cell signaling has been linked to various diseases such as lupus, lymphoma, and various immune disorders. Antigen binding to the BCR promotes the activation of several protein tyrosine kinases (PTK), which leads to phosphorylation of the BCR complex and recruitment and activation of the PTK Syk. This in turn promotes phosphorylation of PLCγ, Shc, and Vav. Additionally, the Tec family member Btk is recruited to the plasma membrane, where it is involved in activation of PLCγ. Initiation of B lymphocyte activation is dependent on the tyrosine phosphorylation-dependent formation of multimolecular effector protein complexes that activate downstream signaling pathways including PKC/Ca2+/NFAT. This cell-based assay has been thoroughly tested and validated by Invitrogen and is suitable for immediate use in screening applications. The following information illustrates the comprehensive assay testing completed and validation of assay performance under optimized conditions (Figure 1).