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S Jahn

Publications and source records attributed to S Jahn.

At least 55 records · Page 3Linked to original sources
PubMed1993

Fine mapping of HIV-1 Nef-epitopes by monoclonal antibodies.

A panel of newly isolated murine monoclonal antibodies is described which are specific for the Nef protein of the human immunodeficiency virus type 1 (HIV-1). Epitope mapping using recombinant Nef-related proteins, synthetic peptides and lipopeptides showed 3 independent antigenic determinants located within the regions of amino acids 83-93, 175-190 and 86-166 of the Nef protein. None of the monoclonal antibodies reacted with recombinant Nef proteins of HIV-2.

Amino Acid Sequence↗
PubMed1993

An anti-lipid A antibody obtained from the human fetal repertoire is encoded by VH6-V lambda 1 genes.

The human hybridoma cell line CB-201 has been obtained from a fusion of fetal spleen lymphocytes (31st gestational week) with heteromyeloma cells. The IgM (lambda) secreted was found to bind to lipid A, whereas other endogenous and exogenous antigens were not recognized. The CB-201 antibody is encoded by the unmutated VH6 gene recombined with DN4 and JH3 elements and a V lambda subgroup 1 gene. Therefore, a VH gene, which was previously described to be over-represented in the fetal repertoire and to be expressed in autoantibody-producing B cells, may encode an anti-bacterial specificity.

Antibodies, Monoclonal↗
PubMed1993

In vitro stimulation of human fetal lymphocytes by mitogens and interleukins.

Human lymphocytes derived from fetal spleen and liver were studied for their capacity to respond to mitogens and interleukins using different in vitro models (cell volume increase, [3H]thymidine incorporation, Ig secretion). Although the number of mature B and T cells in the fetal liver preparations remained nearly constant [Settmacher et al. (1991) Immunobiol. 182, 256], only lymphocytes obtained from fetal organisms before the 25th week of gestation could respond to some of the polyclonal stimulators (PWM, anti-CD3 + IL-2, SAC + IL-2, SAC + IL-4) tested, whereas cells obtained after that period failed. In the fetal spleen, however, with increasing percentages of mature B and T cells during fetal development, a growing ability to respond to mitogens was registered, which, however, did not achieve the values found for the adult spleen material.

Adult↗
PubMed1993

Immune restoration in children after partial splenectomy.

Splenectomy (SE) is recognized to be a therapeutical approach in treating children with severe autoimmune diseases (chronic idiopathic thrombocytopenia; hemolytic anemia) or hypersplenism because of portal hypertension. Nevertheless, removal of a main immune organ results in elevated infection risk for these patients. Partial splenectomy (PSE) was developed as a therapeutical compromise to retain immunologically active spleen tissue. Here, we document the analysis of immune parameters obtained from children after both partial and total splenectomy, which have been followed up for a period of more than 6 years: (i) Lymphocytes from both groups of patients failed to produce IgG in response to pokeweed mitogen in vitro. This was observed in 11/20 splenectomized patients even 10 years after operation, whereas in PSE patients a restoration of this parameter after 1-2 years was seen. (ii) In patients after PSE, but not in splenectomized persons, an elevated number of HLA-class II positive cells had been detected suggesting a different situation of immune regulation following this operation. However, in parallel with an improvement of B cell in vitro activity this parameter was found to achieve normal values. Our findings indicate that partial splenectomy may be a therapeutical alternative, if the therapeutic goal can be achieved by this procedure.

Adolescent↗
PubMed1992

Expression of the HIV-1 Nef protein in the baculovirus system: investigation of anti-Nef antibodies response in human sera and subcellular localization of Nef.

The nef gene of HIV-1 was expressed in insect cells using the eucaryotic baculovirus system. The recombinant Nef protein frequently reacted with seropositive sera of HIV-1 and HIV-2 infected patients. Anti-Nef antibodies in HIV-1 seronegative high risk groups individuals were only occasionally seen. Confocal laser scanning microscopy demonstrated that Nef is present both in the cytoplasm and in the nucleus, indicating that Nef might directly function on gene expression.

Baculoviridae↗
PubMed1992

Oligonucleotide fingerprinting as a means to identify and survey long-term cultured B cell hybridomas and T cell lines.

Common problems encountered during cell culture are cross-contamination, instability, and inadvertent exchange of cells. Here we report on the application of oligonucleotide fingerprinting as a simple and efficient method to screen hybridomas and T cell lines. Among the fingerprint probes tested, the simple repetitive oligonucleotide (CAC)5/(GTG)5 proved to be most useful for obtaining many fragments specific for each cell line. Because of variable loss of chromosomes, cloned hybridoma cells from one fusion exhibit different fingerprint patterns. Thus, antibody-secreting B cell hybridomas can be distinguished easily even when they originate from the same fusion. Furthermore, we were able to monitor the genomic integrity of myelomas and hybridomas over a period of more than 2 years, thereby highlighting long-term stability. Another application of the method is the control of T cell lines requiring irradiated or mitomycin-treated feeder cells for continuous growth or cloning. There cell lines are always threatened to be overgrown by feeder cells that have escaped from the lethal pretreatment. T cell clones of one individual, known to display differently rearranged T cell receptors, did not show differences in their fingerprint patterns. However, in EBV-transformed cloned B cells, slight differences between clones from the same donor were identified.

B-Lymphocytes↗
PubMed1992

Lymphocyte surface marker expression on hybridomas secreting human monoclonal antibodies.

The expression of human leucocyte markers on the surface of hybridoma cell lines producing human monoclonal antibodies was studied using immunofluorescence analysis (FACS). We tested 36 different hybridoma cell lines from fusions of lymphocytes of different organs of fetal and adult organisms with the mouse myeloma line P3 X63 Ag8.653 or the mouse-human heteromyeloma line CB-F7 (IgM-, IgG-, and nonproducer) with a panel of 21 murine monoclonal antibodies against human differentiation and activation antigens. CD2, 3, 4, 5, 8, 10, 23, 25 antigen and major histocompatibility complex (MHC) class II determinants could not be detected on all hybridomas analyzed. The antigens CD22, 69, 71, and 72 were expressed on few of the hybridomas tested. The majority of the cell lines carried the surface markers CD19, 20, 40, 45 as well as the plasma cell markers CD38 and O/C11. The activation antigen 4F2 was expressed on all the cell lines tested. However, a direct connection between the expression of a lymphocyte marker and the capacity for Ig production (high and low producer; Ig isotype), the origin of the lymphocytes, and the fusion cell line used could not be detected.

Antibodies, Monoclonal↗
PubMed1991

A solid-phase enzyme immunoassay for the detection of tetanus toxin using human and murine monoclonal antibodies.

Three human and three murine monoclonal antibodies were tested for their reactivity to tetanus toxin and toxoid and used to establish an enzyme immunoassay specific for tetanus toxin. The dissociation constants of the monoclonal antibodies were between 3.91 x 10(-9) and 8.48 x 10(-12). Two human monoclonal antibodies recognized conformation determinants on the toxin, whereas the others reacted to the heavy chain. Only a combination of antibodies of the two species allowed the development of an enzyme immunoassay for the detection of tetanus toxin with a lower detection limit of 1.2 micrograms/l.

Animals↗
PubMed1991

Characterization of human lymphocytes separated from fetal liver and spleen at different stages of ontogeny.

Membrane markers on human lymphocytes separated from fetal liver and spleen were studied. Depending on the period of intrauterine development, a growing percentage of T- and B-lymphocytes (up to 16% and 45%, respectively) among spleen cells was seen, but in liver the number was low independent of the gestational age (T cells less than 10% and B cells less than 15%). The majority of early CD3+ spleen cells (21st-28th week) expressed TCR alpha beta but not TCR gamma delta, although a significant proportion of these cells was still lacking CD4, CD8, and CD5 differentiation antigens, suggesting their immaturity. Later spleen T cells (29th-36th week) expressed the phenotype as mature adult-type T cells (CD3+TCR alpha beta +CD4/8+CD5+). During ontogeny in fetal spleen, a growing number of B cells could be estimated without any changes in the proportion of subsets, expressing the different light and heavy chains. However, the proportion of CD5+ B cells decreased with gestational age. The results suggest that the functional immaturity of antenatal splenocytes could not be caused by dramatic phenotypical differences in comparison with adult-type splenic lymphocytes.

Antigens, Surface↗
PubMed1991

Human hybridomas derived from CD5+ B lymphocytes of patients with chronic lymphocytic leukemia (B-CLL) produce multi-specific natural IgM (kappa) antibodies.

Great numbers of CD5+ B lymphocytes were detected in the peripheral blood of patients with B-CLL. To study the antibody repertoire of this immune cell subpopulation on a monoclonal level, we fused the lymphocytes derived from five different donors to a highly efficient HAT-sensitive heteromyeloma line (CB-F7). A fusion frequency of up to 10(-5) allowed us to analyse hundreds of initial hybridoma lines per fusion. In all culture supernatants in three out of five fusions IgM lambda antibodies were detected, in two experiments only IgM kappa was measured, suggesting monoclonality of the primary hybridoma cell lines. The later fusions resulted in hybridomas producing multi-specific antibodies against both an autoantigen and an infectious agent: (i) dsDNA/influenza virus haemagglutinin; (ii) dsDNA/class V outer membrane protein type C from Neisseria meningitidis. However, no antibodies of the described specificity were detected in blood sera of patients, indicating a 'switch-on' of the immunoglobulin secretion capacity of malignant B cells during fusion to a myeloma partner. We discuss the results as further evidence for the natural multi-reactive antibody repertoire of CD5+ B cells.

Adult↗
PubMed1991

IgG human monoclonal anti-DNA autoantibodies from patients with systemic lupus erythematosus.

We describe the production of six mouse-human heterohybridomas secreting human IgG anti-dsDNA antibodies derived from patients with systemic lupus erythematosus (SLE). Peripheral blood cells used for fusion experiments were from patients who were shown to have high numbers of anti-DNA secreting B cells in the peripheral blood. All monoclonal antibodies bind to dsDNA in ELISA systems, five are reactive with Crithidia lucilae kinetoplasts and three precipitate dsDNA in the Farr assay. Inhibition studies revealed a remarkable specificity for certain polynucleotide structures. To our knowledge these are the first hybridomas described in the human system that secrete anti-dsDNA antibodies of the IgG class.

Antibodies, Antinuclear↗
PubMed1991

Ultrastructural localization of gold-labeled anti-IgM-antibodies and type A retrovirus particles in endoplasmic cisternae and vesicles of IgM-producing human B cell hybridomas.

Simultaneous appearance of IgM and type A retro-virus particles in endoplasmic cisternae and vesicles of human hybridomas has been demonstrated for the first time by immunogold-labeling at ultrastructural level. A suspected link between type A particle and IgM gene-expression in hybridoma cells could not be substantiated in this study.

Animals↗
PubMed1990

Production and characterization of monoclonal antibodies against human Cu/Zn superoxide dismutase and the establishment of a super-rapid enzyme-linked immunosorbent assay (SURALISA).

Murine monoclonal IgG1 antibodies directed against four different epitopes of human Cu/Zn superoxide dismutase (SOD) were produced by immunization with recombinant Cu/Zn SOD. The antibodies reacted well with the recombinant protein and Cu/Zn SOD purified from human erythrocytes, with binding constants ranging from 8.8 X 10(9) to 2.2 X 10(10) l/mol. When mixed, these antibodies completely prevented the binding of rabbit and sheep polyclonal antibodies raised against erythrocyte Cu/Zn SOD. Whereas one antibody was directed against a common homology region of bovine and human Cu/Zn SOD, all the other antibodies reacted exclusively or preferentially with human Cu/Zn SOD. Only one epitope on the human Cu/Zn SOD molecule was accessible at two different sites as demonstrated in a homologous two-site assay with one and the same antibody used as both capture and indicator antibody. In the indirect two-site assay with unlabelled monoclonal antibodies, and additive effect with a steeper dose-response curve was obtained by mixing antibodies against different epitopes. A super-rapid one-step two-site enzyme immunoassay (overall duration 20 min) was established with antibodies against two different epitopes. Its detection limit was 0.5 micrograms SOD/l.

Antibodies, Monoclonal↗
PubMed1990

Immunochemical quantification of Cu/Zn superoxide dismutase in prenatal diagnosis of Down's syndrome.

Cu/Zn superoxide dismutase (SOD) was quantified by enzyme immunoassay for prenatal diagnosis of Down's syndrome. Overall, 154 samples of amniotic fluid, 72 samples of amniotic cells and 31 samples of chorionic tissue were investigated. Due to the large biological variance of the SOD concentrations in normal pregnancies (range for amniotic fluid 10.5-154.9, for amniotic cells 40.0-338.8, and for chorionic tissue 132.2-649.5 g SOD/g protein) the cases of Down's syndrome detected by karyotype analysis were not reliably identified by Cu/Zn SOD quantification. As in erythrocytes obtained from patients with Down's syndrome, a trisomy 21 was easily and accurately detected in the erythrocytes from very small quantities (about 50 microliters) of umbilical blood. The SOD concentrations in normal cases (n = 40) varied between 11.4 and 17.3 and in the cases of trisomy 21, as confirmed by karyotyping (n = 4), between 22.5 and 23.2 ng/one million cells. SOD quantification in fetal erythrocyte is a helpful additional method in prenatal Down syndrome diagnosis under certain conditions, which are discussed.

Amniotic Fluid↗
PubMed1990

Polyclonal stimulation of human B lymphocytes derived from fetal liver and spleen cells at different stages of ontogeny.

The functional capacity of human lymphocytes derived from fetal liver and spleen at different stages of ontogeny (16-34 weeks of gestation) was studied using in vitro models (increase in cell volume, [3H]thymidine incorporation, Ig secretion) reflecting various stages of activation induced by mitogens (LPS, PWM) in vitro. Lymphocytes differed in their reactivity to LPS depending on the period of intrauterine development: cells from the early liver could respond with enhanced IgM production whereas lymphocytes derived from this organ after more than 25 weeks failed. The opposite was found to apply to spleen cells: only lymphocytes derived from the organ after more than 25 weeks showed significant LPS-induced in vitro differentiation. These data were in correlation with the proliferative response to LPS. It was clear that the number of CD20-positive mature B cells in the lymphocyte preparations was not responsible for these results, since comparable yields were found throughout the period of fetal development of the liver studied, whereas in the spleen increasing numbers of B cells were seen.

B-Lymphocytes↗
PubMed1990

Immortalization of magnetically separated human lymphocytes by electrofusion.

Human lymphocytes from peripheral blood (MNC) were separated on magnetic beads for the presence of different surface markers. Cells from positive and negative fractions were successfully immortalized by electrofusion with the heteromyeloma line CB-Fu2. B cells, which were separated on anti-CD 19 coated beads, could be immortalized at a rate between 10(-5) and 10(-4) even if the fusion was conducted with just a few hundred thousand cells. Comparison of the frequency of Ig-positive hybridomas in B cell, T cell, and unseparated MNC fusions indicated that also non-B cells may give rise to HAT resistant hybridoma clones, although the fusion frequency was low.

Animals↗
PubMed1990

The hybridization of EBV-immortalized human B-lymphocytes with a human-mouse heteromyeloma cell line.

The combination of Epstein-Barr-Virus (EBV)-permitted immortalization and somatic hybridization (fusion with a myeloma partner) may be the method of choice to produce human monoclonal antibodies. We show here that the fusion of EBV-infected human B-lymphocytes to the HAT-sensitive, ouabain-resistent heteromyeloma (human x mouse) fusion line CB-F7, resulted in stable growing hybridomas producing much more immunoglobulin than the parental lymphoblastoid lines. A more efficient clonability was shown for hybridoma cultures too. The loss of B cell markers (HLA-class II antigen, CD-22, CD-37) was detected. Limiting dilution experiments showed a better fusionability of IgM-producing EBV-transformed B cells in comparison to IgG-secreting counterparts.

Animals↗