This chapter provides information on the application of flow cytometry for analysis of B-cell development, describing in detail the particular surface proteins that can serve as markers for recognizing distinct stages in this process. These cell fractions range from just prior to initial heavy chain rearrangement, the germline pro-B stage, through D-J rearranged pro-B and heavy chain expressing pre-B stages, to the maturing surface BCR positive B-cell stages. It also outlines assays for the characterization of these cells, including procedures for testing functional lineage restriction, determination of rearrangement status, analyses of gene expression at the ribonucleic acid (RNA) and protein level, and assessment of cell cycle state.