Membrane-bound Ig were studied by immunofluorescence methods in 116 patients with lymphoproliferative disorders. Surface Ig synthetized in vitro were studied in many patients. In 70 of 73 cases of chronic lymphocytic leukemia (49 without and 24 with monoclonal serum Ig), monoclonal surface Ig with a distribution for heavy and light chains similar to that of normal lymphocytes were found. In three cases, these surface Ig accumulated as crystals in the lymphocyte cytoplasm. Studies limited to staining of freshly drawn cells may lead to erroneous conclusions since serum antibodies may become bound to the cell surface and simultaneous positivity for µ, γ, κ, and λ, due either to the attachment of immune complexes at the lymphocyte surface or to anti-IgG antibody activity of the monoclonal surface IgM, was not rare. IgM with anti-IgG activity was found on the surface of both lymphocytes and lymphoblasts in a patient with acute transformation of chronic lymphocytic leukemia. In a single case, µ and γ chains were simultaneously found on some lymphocytes besides two single producer clones. A biclonal proliferation characterized by distinct surface Ig markers was demonstrated in three other cases of lymphocytic leukemia and in a patient with Waldenström’s macroglobulinemia. In this latter condition (31 cases), most marrow lymphoid cells including plasma cells and the majority of blood lymphocytes carried IgM determinants which had the same light chain type as the serum IgM and which shared its eventual antibody activity. Similar results were obtained in a few patients with the hematological features of macroglobulinemia but with serum monoclonal IgG or IgA or with unreleased IgM. Studies on cases of γ and α heavy chain diseases, cold agglutinin disease, leukemic reticuloendotheliosis, and Sezary’s reticulosis are also recorded. The value of surface Ig as B cell markers in lymphoproliferative diseases is outlined.

Lymphocytes from a patient with Waldenstrom's macroglobulinemia and known anti-immunoglobulin IgG activity of serum monoclonal IgM and lymphocytes from selected patients with chronic lymphocytic leukemia were studied by the membrane immunofluorescence procedure. Freshly drawn lymphocytes were shown to bear simultaneously i, y, x, and X chain determinants. Experiments combining redistribution induced by antibody and double labeling proved that IgG was bound to surface IgM. After removal of surface immunoglobulins by treatment with trypsin followed by incubation in culture medium, or after redistribution induced by antibody, the exclusive presence of a newly synthetized monoclonal IgM was demonstrated. Several experiments showed that this surface IgM does specifically bind normal human IgG molecules devoid of aggregated material. The IgG molecules could be removed from the cell surface by lowering the pH. In addition to its high incidence among serum monoclonal macroglobulins, anti-IgG activity of membrane-bound monoclonal IgM is not uncommon in patients with chronic lymphocytic leukemia, a disease that provides homogeneous populations of lymphocytes derived from bone marrow, with receptor sites of defined antibody activity.The presence of receptor molecules of immunoglobulin on the surface of lymphocytes derived from bone marrow (B lymphocytes) is well documented in different species including man (1, 2). Since lymphocytes from many patients with chronic lymphocytic leukemia (1, 3, 4) and from patients with Waldenstr6m's macroglobulinemia (5) are characterized by a monoclonal surface IgM marker, these immunoproliferative disorders may provide suitable models for the study of antibody activities of surface immunoglobulins on homogeneous B-lymphocyte populations. In view of the high incidence of antibody activity against IgG among serum monoclonal IgM (6), it appeared logical to search for such an antibody activity of surface monoclonal IgM. We have studied the cells of a patient with Waldenstr6m's macroglobulinemia and known rheumatoid-factor activity of serum monoclonal IgM. Moreover, we have found by immunofluorescence study of living cells the simultaneous presence of Sy-, x-, X-, and usually u-chain determinants on the lymphocytes from several patients with chronic lymphocytic leukemia. One of the possible explanations for this unexpected finding is the anti-IgG activity of surface immunoglobulins. The results presented here clearly show that, in some instances, the monoclonal IgM on the surface of lymphocytes does specifically bind normal human IgG. MATERIALS AND METHODSThe methods used for isolation of lymphoid cells from blood or bone marrow, immunofluorescence study of living cells in suspension, and preparation and characterization of reagents stained with fluorescein or rhodamine that are monospecific for y, lu a, x, and X chains have been described (4, 5). Pooled normal human IgG was purified by chromatography on diethylaminoethyl (DEAE)-cellulose columns and coupled to tetramethylrhodamine isot...

Idiotype-bearing and antigen-binding receptors produced by blood T lymphocytes in a case of human myeloma"Twenty-five t o 30 % of blood lymphocytes from a myeloma patient with an IgGl(K) protein with antibody activity to horse a2-macroglobulin (a2M) reacted with purified IgG from a rabbit antiserum t o the idiotypic determinants of this protein. A small proportion of these idiotype-bearing lymphccytes were IgG-bearing B cells, but most of them were T cells. Several experiments demonstrated the actual synthesis of the idiotypic structures carried by the T lymphocytes. All idiotype-bearing lymphocytes bound horse a2M, and the structures that bore the idiotype and bound the antigen were shown t o completely cocap. After biosynthetic labeling with I4C-labeled amino acids, immunoprecipitation and sodium dodecyl sulfate gel electrophoresis, the antiidiotypic serum and the a2M antigen were found t o react with the same molecules. Preliminary data & the size of these antigen-binding T lymphocytederived molecules are in accord with those of Binz and Wigzell ( S c u d . ; I . Immunol. 1976. 5: 559) in the rat. The finding in this patient with multiple myeloma of a homogeneous and possibly malignant population of T lymphocytes, synthesizing identical antigen receptors which share idiotypic (but not isotypic) determinants of the immunoglobulin molecule produced by the malignant B cell clone, therefore confirms in man the results of several recent studies in other mammalian species. Abbreviations: PBL: Peripheral blood lymphocytes IBL: Idiotypebearidg lymphocytes; lymphocytes bearing (and synthesizing) the same idiotypic determinants as those of the serum myeloma protein SIg: Surface immunoglobulins a2M: ~~2-Macroglobulin RFC: Rosette-forming cells SDS: Sodium dodecyl sulfateThe presence, in human and murine myelomas, of peripheral blood lymphocytes (PBL) bearing on their membrane molecules which react with antisera specific for the individual myeloma protein idiotypic determinants, has been reported by several groups [l-41. The fact that these molecules were actual cell products was demonstrated by their regrowth after stripping of the membrane by proteolytic enzymes [3-51.Blood lymphocytes which carry the idiotypic determinants of the myeloma protein, here referred t o as "idiotype-bearing lymphocytes" (IBL), were considered t o be B cells. This interpretation is supported by our finding, in about one third of untreated myeloma patients, of "monoclonal" populations

H L A antigens and membranebound immunoglobulins 297 ral antibodies are elicited is not unique. Benjamini and his collaborators (personal communication) have observed a similar phenomenon when testing the cross-reaction between native lysozyme and its reduced carboxymethylated derivative; similarly, Michaeli* and his colleagues have encountered cellmediated cross-reaction between Ascaris and human collagens, whereas n o humoral crossreactivity between them could be detected. It seems, therefore, that the use of cell-mediated immune response may be o f help in following phylogenetic relationships between proteins which might have diversified during evolution t o such an extent as t o prevent humoral cross-reac tion.The financial support of the Ford Foundation is gratefully acknowledged.