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Studies on lymphocyte subpopulations and NK cell activities in epileptic patients.

SummaryThe subpopulations of T lymphocytes, B lymphocytes and macrophages of the peripheral blood in 58 cases of epilepsia were studied with McAb by IFA method. The activity of natural killer (NK) cells of penpheric blood monocyte was investigated by51Cr-releasing method. 22 healthy donors of about the same age served as controls. The results showed that the percentage of T3 and T4 lymphocytes in epileptic patients was decreased, and the ratio of T4/T8 was markedly reduced as compared to control group. The percentage of T8 lymphecytes was increased. There were no significant changes in B lymphocytes and macrophages. The NK cell activity also showed decrease. The results suggested abnormality in cellular immunity in epilepsia, which may be involved in the pathogenetic mechanism of the disease.

Studies on monoclonal anti-idiotypic antibodies to human B cell leukemia.

SummaryMurine immunocytoma cell line, NS-1, was fused with spleen cells of Balb/C mice which had been stimulated by tolerogenic disaggregated human gamma globulin and immunized by purified serum IgM from the patient with chronic B cell leukemia (B-CLL). 10 hybridoma cell lines secreting monoclonal anti-idiotype (anti-Id) antibodies to human CLL were obtained. The McAbs were subclasses belonging to IgM and of IgG mouse. Specificity and biologic characters of the monoclonal anti-Id antibodies from culture fluid or ascites were assayed by ELISA, indirect mixed ELISA sandwich, ELISA inhibition, immunofluorescence (IF) and IF inhibition. The study also proved that monoclonal anti-Id antibodies could react with homologous IgM, but not with Ig from normal donors or a panel of patients with myeloma. The results of IF and IF inhibition assay showed that monoclonal anti-Id antibodies were bound to lymphocytes of patient with B-CLL. Their reactivity was inhibited by homologous IgM, but not by lymphocytes of patients with ALL or lymphoma. Monoclonal anti-Id antibodies were heterogenous reactive patterns with cell lines in vitro.

Enhancement of B cell responses by the interaction of CD2 with LFA-3

SummaryPotential effect of the interaction of CD2 with its ligand LFA-3 on B cell responses in vitro was studied to evaluate the action of CD2-LFA-3 in immune responses. It was found that: 1) addition of autologous erythrocytes to uniractional mononuclear cells resulted in enhancement of PWM-induced lymphocyte proliferation, and had no significant influence on non-mitogen stimulation; 2) autologous erythrocytes potentiated the synthesis of immunoglo bulins; and 3) costimulating effects of autologous RBC can be depressed by preincubation of PBMNC with anti-CD2 monoclonal antibody (WuTll). The results suggest that preincubation of mononuclear cells with anti-CD2 McAb has a down-regulating effects on B cell responses and the interaction of CD2 with its ligand LFA-3 expressed by RBC is closely related to immune responses.

Studies on monoclonal anti-isotypic and anti-idiotypic antibodies against leukemia and myeloma: I. preparation and characteristics of monoclonal anti-isotypic antibodies to igm from B chronic lymphocytic leukemia

SummaryFive hybridomas producing McAbs against human serum IgM-isotype were obtained by fusing the myeloma NS-1 cell and the RALB/C murine spleen cell which had been immunized by serum IgM of patient with B chronic lymphocytic leukemia (B-CLL). These McAbs only reacted with human IgM, not with other immunoglobulins in ELISA and immune double diffusion test. An approximate positive rate of peripheral blood lymphocyte (PBL) was got when these McAbs and the McAbs aginst B cell were tested by indirecl immunofluorescent assay. The positive rate was similar to that obtained by direct immunofluorescent test. Immunoblotting showed that the molecular weight of the antigen to these McAbs was 70–90 Kd, indicating that the antigen was IgM. The practical value of the McAbs against human IgM was also discussed.

Studies on monoclonal anti-isotypic and anti-idiotypic antibodies against leukemia and myeloma: I. study on monoclonal anti-idiotypic antibodies against λ chain protein of myeloma

SummaryWe used A chain extracted from urine of patient with myeloma (IgD λ) as antigen for immunizing BALB/C mice, and 86 hybridoma cell clones secreting monoclonal antibodies (McAb) were obtained after fusing twice and cloning 3–4 times. 15 of these clones secreted monoclonal anti-idiotypic antibodies (anti-Id McAb). The results showed that 12 of 15 anti-Id antibodies reacted only with homologous λ chain and IgD, not with A chain, κ chain, IgG, IgA, IgM, IgD, IgE, albumin and paraglobulin from normal subjects. Indirect immunofluorescent assay demonstrated that positive reaction rate between anti-Id McAbs and peripheral blood lymphocytes or bone marrow cells of the patient with myeloma was up to 23 %. No reaction was observed between anti-Id McAb and peripheral blood lymphocytes or bone marrow cells from normal subjects. Some of these McAbs presented positive reaction with plasmacytoma cell lines cultured in our laboratory.

Immunomodulation of musk-moxa-string therapy in patients with scrofula.

SummaryThe effects of musk-moxa-string therapy on the immune system in man were investigated in 39 patients with scrofula. Before treatment, the numbers of peripheral blood (PB) CD3+ and DC4+ cells and the ratio CD4+/ CD8+ were found to be lower in patients with scrofula than in normal subjects, while those of B cells and DR+ cells were higher. Response of peripheral blood mononuclear cells (PBMC) to phytohemagglutinin (PHA) diminished in patients with scrofula. At month 2–6 of musk-moxa-string therapy the number of PB CD8+ cells showed slight diminution along with significant increaes in CD3+ and CD4+ cells and CD4+/CD8+ ratio in total lymphocytes (P<0.001). In vitro a marked increased blastogenic response to mitogen stimulation with PHA was observed in PBMC of patients with scrofula after treatment (P<0.001). In contrast, B lymphocytes, monocytes, DR+ cells and blastogenic response to concanavalin A and pokeweed mitogen were not influenced by musk-moxa-string therapy.

Studies on monoclonal anti-isotypic and anti-idiotypic antibodies against leukemia and myeloma: VI. Purification and relative affinity of monoclonal antibodies

SummaryIn this study, the anti-idiotypic and anti-isotypic antibodies (McAbs) against IgM of the Patient with B chronic lymphocytic leukemia (B-CLL) were purified from hybridoma ascites by n-Octoic acid precipitation method. The purified McAbs have high purifity and high antibody activity as evidenced by immunoelectrophoresis, SDS-PAGE and ELISA. Relative affinity of 11 McAbs was measured by using indirect ELISA and double antibody sandwich ELISA method. It was found that relative affinity of various McAbs to the same antigen was different. 11 McAbs could be divided into two groups by analysing their 50 % maximum binding. The relative affinity of 4 McAbs in the culture supernatants was consistent with that of McAbs in the purified ascites. Our experimental results provide an important basis for rational application of these McAbs.

Studies on monoclonal anti-isotypic and anti-idiotypic antibodies against leukemia and myeloma: IV modulation of membrane fluidity of leukemic cell lines and tonsillar cell stimulated with McAbs

SummaryIn this study the technique of labelling the cell membrane with DPH fluorescence polarization was used to observe the membrane fluidity of B lymphocytic cell lines and tonsillar cells from healthy persons; the modulation effect on membrane-fluidity induced by McAbs against isotypic and idiotypic determinants of IgM from patients with leukemia was studied as well. The expression of the corresponding isotypic and idiotypic determinants of IgM on the cell membrane was determined. The results show that the membrane fluidity of leukemic cell lines is remarkably higher than that of tonsillar cells from healthy persons, and McAbs against isotypic determinants of leukemic IgM can enhance the membrane fluidity of all kinds of cells mentioned above. However, the anti-idiotypic monoclonal antibody increased only the membrane fluidity of leukemic cell lines. These results indicated that there was a close relationship between the effect of McAbs on cell membrane fluidity and the expression of corresponding isotypic and idiotypic determinants of IgM on the cell membrane.

Studies on monoclonal anti-isotypic and anti-idiotypic antibodies against leukemia and myeloma: V. The effects of monoclonal antibodies and interferon on the levels of cyclic nucleotides in leukemic cell lines.

SummaryAfter the leukemic cell lines were treated with monoclonal antibodies (McAbs) and interferon (IFN-α), the changes of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) levels in the corresponding leukemic cell lines were measured by radioimmunoassay. The results showed that when the ratio of antigen to antibody was 80 to 1, the cAMP levels in the leukemic cell lines were obviously higher than those in the controls while the cGMP levels were obviously lower after being treated with the corresponding McAbs for 16–24 h (P<0.001). The average level of intracellular cAMP was remarkably increased and that of cGMP underwent no significant changes in the leukemic cell lines after treatment with IFN-α.

ANALYSIS ON EPITOPES OF IGM WITH MONOCLONAL ANTI-ISOTYPIC AND ANTI-IDIOTYPIC ANTIBODIES AGAINST IgM FROM B-CLL

A double antibodies additivity ELISA test was employed to identify the epitopes which can be recognized by monoclonal antibodies (McAbs) against IgM from B chronic lymphocyte leukemia (B-CLL). The computer grouping programme analysis showed that 4 anti- isotypic MaAbs could be divided into two groups and 10 anti- idiotype McAbs could be divided into four groups. The result was consistent with that of the indirect sandwich ELISA and inhibition ELISA test. It suggested that there were at least 6 distinct IgM epitopes which can react specifically with 14 McAbs. Our study indicated that the combination of the additivity ELISA test and the computer grouping programme analysis is of help in studying the relationship of the structure and function of antigen.