B-cell chronic lymphocytic leukemia (B-CLL) is characterized by highly variable distribution of tumor mass between peripheral blood, bone marrow and lymphoid organs which is important for staging, classification and prognosis. These clinical findings with novel data about importance of B-cell receptor and its stimulation with the support of microenvironment indicate important role of tissues (lymphoid organs and bone marrow) in the pathogenesis of B-CLL. Here is presented the novel approach of simultaneous characterization of B-CLL cells form peripheral blood, bone marrow and lymph nodes by flow cytometry and immunocytochemistry, defining inter- and intraclonal diversity with respect to various molecules. These include adhesion molecules (integrins, immunoglobulins, selectins), chemokine receptors (including CXCR-4), signaling molecules and prognostic factors (CD38 and ZAP-70), proliferation and apoptosis markers (including Ki67, AgNORs with PK index, survivin, bcl-2) and therapeutic targets (CD20 and CD52) and residual hematopoietic stem cells. A number of interesting significant interactions have been discovered, pointing to the important role of neoplastic cell microenvironment. These may in addition to insights in pathogenesis and roles of different microenvironments add to diagnosis, prognosis and treatment of B-CLL patients.