We studied 61 CD20- B-cell lymphomas, including 29 cases of precursor B-cell lymphoblastic leukemia/lymphoblastic lymphoma (B-ALL/B-LBL), 25 cases of CD20- recurrent mature B-cell lymphoma after rituximab therapy, and 7 cases of CD20- diffuse large B cell lymphoma (DLBCL). We used markers specific for B lineage: CD79a, Pax-5, OCT.2, and BOB.1. All B-ALL/B-LBLs expressed Pax-5 (29/29 [100%]), 25 (93%) of 27 expressed BOB.1, 23 (79%) of 29 expressed CD79a, and 6 (22%) of 27 expressed OCT.2. The percentages of cases expressing Pax-5, CD79a, OCT.2, and BOB.1 in CD20- recurrent mature B-cell lymphomas after rituximab treatment were 88% (21/24), 84% (21/25), 81% (17/21), and 73% (16/22), respectively. CD20- DLBCLs rarely express routine B-lineage markers, such as and CD79a and Pax-5, but they expressed OCT.2 or BOB.1. Pax-5, BOB.1, and CD79a antigens are the most reliable B-lineage markers for paraffin immunophenotyping B-ALL/B-LBL. CD79a and Pax-5 should be used as the first-line B lineage-specific markers for rituximab-treated CD20- mature B-cell lymphomas. If negative, OCT.2 or BOB.1 may be useful. The newly identified B-lineage markers, OCT.2 and BOB.1, may be the most useful for the B-lineage determination of CD20- plasmablastic or primary effusion subtypes of DLBCL.