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Biomedical subjects

S Jahn

Publications and source records attributed to S Jahn.

At least 73 records · Page 4Linked to original sources
PubMed1990

[Structure and function of the immunoglobulin Vh gene in man].

The present study provides a survey on the structure and functions of human Vh-genes. The more than 100 individual gene segments on chromosome 14 are classified, according to sequence homologies, into 6 families. They differ very much in size and contain pseudogenes, form an interspersed cluster, exhibit homologies with mouse genes and have phylogenetically developed from a single primordial gene. There seem to be no association of antibody specificities with distinct Vh genes or families. Reports of preferential VH gene usage in the fetal stage or in autoantibody synthesis require confirmation. Vh genes for Ig production in the adult mammalian organism are, most probably randomly selected.

Antibody Specificity↗
PubMed1990

Multireactive human monoclonal antibodies.

Human monoclonal antibodies were tested using different immunochemical procedures for their reactivity with various antigens. The great majority of human monoclonal IgM antibodies (15 out of 24) turned out to bind to a whole series of recognized antigens (DNA, keratin, tetanus toxin, ricin etc.). The specificity of these reactions was detected by competitive assays. For some antibodies a simultaneous reaction with two antigens could be demonstrated by capture bridge technique. IgG antibodies also turned out to be multireactive, but not to the same extent (range of antigens much smaller, only 5-6 out of 53).

Antibodies, Antinuclear↗
PubMed1990

Strategies in the development of human monoclonal antibodies.

A high-efficiency, HAT-sensitive heteromyeloma fusion line CB-F7 has been developed from an 8-Azaguanin-treated Ig-non-secreting human X mouse heterohybridoma. The use of this line allowed us to produce human hybridomas more successfully by fusion of cell material from blood, lymph node or spleen. A polyspecific repertoire of IgM isotype was detected among the hybridomas obtained from the spleen. These IgM antibodies reacted with autoantigens as well as with foreign material. This naturally occurring repertoire may be of interest since it has anti-bacterial activity. The frequency of the occurrence of polyspecific antibody-producing hybridomas was high in the spleen. Apart from the detection of polyspecific IgM antibodies we did not find IgG-secreting hybridomas with anti-bacterial reactivity among thousands of initial lines derived from non-immunized persons. We therefore tried to fuse lymphocytes from donors, who were boosted with Tetanus Toxoid (TTd). A short and limited optimum time period at which the blood should be taken from the donors after boosting, was detected. Seven days after in vivo immunization high yields of IgG-anti-TTd-producing lines (239 out of 731 IgG-producers) were found. The methods of developing more efficient production of human monoclonal antibodies of a pre-defined specificity were discussed.

Animals↗
PubMed1989

[Chemical characterization of Escherichia coli K 1 capsular polysaccharide].

The exact chemical characterization of the prepared Escherichia coli K 1 capsule polysaccharide is necessary and a prerequisite for using this antigen as a screening antigen in the production of monoclonal anti-K 1-antibodies. The K 1-antigen, prepared by phenol-water-extraction, was analyzed by protein, RNA, and neuraminic acid determination. An addition, the antigen was subjected to elementary analysis, infrared- and 13C-NMR-spectroscopy, and gelchromatography. After testing the serological specificity, the K 1-antigen was identified as a high molecular polyneuraminic acid.

Antigens, Bacterial↗
PubMed1989

Development of specific IgG-producing human hybridomas by fusions of lymphocytes from actively immunized persons.

More than 13,500 initial hybridoma lines derived from fusions of lymphocytes from non-boosted persons were tested for IgG production against Tetanus Toxoid. However, only 2 were found to produce IgG monoclonal antibodies of the desired specificity. Peripheral blood lymphocytes were then taken from actively immunized donors 3, 7, 14 or 60 days after boosting and fused to the HAT-sensitive heteromyeloma cell line CB-F7. A strong enhancement of both IgG-producing hybridomas and specific IgG-producers was detected in fusions of lymphocyte material derived 7 days after booster injection (239 of 731 IgG-producing lines showed anti-TT-specificity). Two months after boosting no more specific IgG-producing hybridomas could be established from the peripheral blood of the donors. Data were discussed regarding an optimized human monoclonal antibody production against bacterial antigens.

Antibodies, Bacterial↗
PubMed1989

Occurrence of hybridomas producing multispecific IgM in human x (human-mouse) fusions with lymphocytes from different human immune compartments.

Human lymphocytes derived from peripheral blood, the spleen and lymph nodes were fused to the HAT-sensitive heteromyeloma cell line CB-F7. The Ig-producing initial cell lines were selected, and the supernatants were further analyzed for specific antigen binding (ELISA). IgM-antibodies were found which reacted with self- and non-self antigens of different molecular origin (nucleotides, proteins, carbohydrate structures). These antibodies were called multireactive (multispecific). The multispecific IgM-producing human hybridomas occurred with higher frequencies in the spleen (6.9% of IgM-producers) cell fusions than in experiments where peripheral blood-derived lymphocytes were fused (2.7%). There were no hybridomas producing multireactive antibodies detected in fusion material from lymph nodes. The greatest number of multireactive IgM was seen when PBL from SLE or anti-HIV-positive patients were hybridized to CB-F7 cells. Representative cell lines were cloned and recloned. The multireactivity of the IgM produced by really monoclonal cells, however, was preserved.

Animals↗
PubMed1989

Development of specific human mab's by a small scale electrofusion technique: the influence of some physical and chemical factors on hybridoma yield of human peripheral blood lymphocytes XCB-F7 fusions.

A fusion chamber and an appropriate procedure are described which allow to fuse a sample of 15 to 25 microliters of cell suspension every two minutes. The cells can be observed throughout the process. They are not exposed to mechanical stress after the fusion pulse. Electrofusion between the heteromyeloma line CB-F7 and human mononuclear cells from peripheral blood of immunized donors is shown to provide stable hybridomas producing IgG against tetanustoxin. Pronase treatment, calmodulin, PEG, lanthanum and a number of variations in the fusion conditions were investigated as to whether they influence physical fusion of the cells, hybridoma yield, and immunoglobulin production.

Animals↗
PubMed1989

Cryopreservation of newly formed human and mouse hybridoma cells.

The aim of this study was to develop an optimal technique for cryopreservation of newly formed human and mouse hybridoma cells immediately after fusion. It was shown that human hybridoma cells could be frozen most successfully at a cooling rate of 5 K/min whereas mouse hybridomas at 1 K/min. The percentage of FCS in cryopreservation media (30 or 90%) had no influence on the recovery of both types of hybridomas. For testing the efficiency of freezing, the fusion rate (number of growing and Ig producing hybridomas), the ratio of IgM:IgG-producing initial hybridoma lines and yields of specific wells were analysed. Comparable results were registered when optimally cryopreserved and non-frozen material was studied. Optimization of the cryopreservation of newly formed hybridomas may be of methodological importance, especially when a time-consuming screening for antigenic specificities must be carried out.

Animals↗
PubMed1988

Establishment of human Ig producing heterohybridomas by fusion of mouse myeloma cells with human lymphocytes derived from peripheral blood, bone marrow, spleen, lymph node, and synovial fluid. Effect of polyclonal prestimulation and cryopreservation.

50 fusion experiments were carried out to analyse heterohybridization efficiencies on mouse myeloma cells of the P3 X63 Ag8/653 line with human lymphocytes derived from peripheral blood, bone marrow, lymph node, spleen or synovial fluid. We found higher yields of growing and human Ig-producing hybridoma lines when lymphocytes from spleen or lymph node were fused. Although primary hybridomas could be established from fusions with bone marrow-derived cells, only in nine out of 1616 initially seeded wells was Ig production registered. Four fusions using immune cells from synovial fluid were made without success. Independently of the source of lymphocytes pokeweed mitogen (PWM) prestimulation had no enhancing effect on the percentage of wells with cell growth and this did not alter the IgM:IgG ratio in primary hybridomas (9:1), although cells from all compartments used here (with the exception of bone marrow cells) could be stimulated with PWM to produce both IgG and IgM in cultures. Cryopreserved lymphocytes from different sources could be used for fusions with comparable results registered for the fresh material.

Animals↗
PubMed1988

The high efficiency, human B cell immortalizing heteromyeloma CB-F7. Production of human monoclonal antibodies to human immunodeficiency virus.

This paper describes the construction of a new heteromyeloma cell line designated CB-F7. The cell line was derived from xenogeneic somatic cell hybridization between normal human B lymphocytes and the murine HAT-sensitive P3X63Ag8/653 cell line. CB-F7 cells were characterized by rapid cell growth (doubling time about 16 h) and high cloning efficiencies in culture medium supplemented with 10% or 5% fetal calf serum, respectively. The karyotype of the cells consists of about 75-78 chromosomes as well as two chromosomal fragments. Fusions of the cells with human peripheral blood cells resulted in approximately 2-6 clones per 10(5) seeded lymphocytes. Furthermore, the cells are ouabain resistant and therefore suitable for fusions with EBV-transformed lymphoblastoid cell lines. Using CB-F7 as the parental cell line a number of specific human mAb producing hybrids were established. For the first time, we describe here the generation of hybrids secreting human monoclonal antibodies to human immunodeficiency virus (HIV). Two monoclonal antibodies of IgG type and one of IgM type reacted with the major core protein p25 and one IgG antibody reacted with the transmembrane protein gp41.

Antibodies, Monoclonal↗
PubMed1988

A myelopeptide from unstimulated bone marrow cells with immunomodulatory activity in lymphocyte cultures from healthy donors and patients with hypogammaglobulinemia and active lupus erythematosus.

A myelopeptide (SAP) was derived from culture supernatants of unstimulated animal bone marrow. SAP consists of a group of peptides with a molecular weight of about 2000 D, having a broad variety of biological activities. Testing immunoregulatory properties of the purified factor Petrov, Mickhailova & Zacharova [(1971). Immunoglobin synthesis in syngenic cells of different lymphoid tissues. J. Immun., 106 1086-1089] found enhanced antibody production in mice (SAP-stimulator of antibody production). We show here that the substance could induce expression of activation markers on human lymphocytes (4F2, HLA-class II antigens, thermostable SE rosette formation) and potentiate their appearance in combination with mitogens (PWM, PHA, Con A). Although SAP was not mitogenic for itself, it enhanced lectin-induced 3H-thymidine incorporation and T-cell-dependent B-cell differentiation in a dose-dependent manner. The factor was able to reconstitute disturbed PWM-driven Ig synthesis in lymphocyte cultures derived from two patients with hypogammaglobulinemia and a healthy non-responder to PWM. On the other side, SAP potentiated the inhibitory activity of PWM on elevated spontaneous IgG secretion in cultures derived from patients with active SLE. Findings of this study indicate immunomodulatory capacity of SAP on human peripheral blood lymphocytes possible via T-cell activation. The results suggest a potential therapeutic application of SAP in patients with disturbances in the T-dependent B-cell differentiation.

Agammaglobulinemia↗
PubMed1988

Partial splenectomy for treatment of chronic idiopathic thrombocytopenia in childhood.

The aim of this paper was to demonstrate the efficiency of partial splenic resection in treatment of chronic idiopathic thrombocytepenie purpura (Werlhof-Disease). In all but one case remissions of thrombocyte counts over 100 Gpt/l could be achieved. Immunological studies demonstrate the existence of a suppressor-state after this operation, which could be responsible for success of this procedure. In contrast to general considerations the partial-splenectomized child must be considered as an immunological deficient organism for about 6 to 12 months. Restoration of immunological-means whole splenic functions-parameters is to expect always after this time.

Adolescent↗
PubMed1988

Feeder cells from different sources and conditioned media for recloning of human--mouse and mouse--mouse hybridomas.

The use of different feeder cell layers (peritoneal macrophages from mice and rats, spleen cells and thymocytes from mice) for recloning of human--mouse and mouse--mouse hybridomas has been described. Optimal numbers of feeder cells from different sources required for high cloning efficiencies were determined. It was possible to use cryopreserved rat and mouse macrophages as feeders for cell cloning. However, the resulting cloning efficiency was much lower in comparison to the fresh material. Culture supernatants from human endothelial cells (added in a final concentration of 40% v/v) and from chicken embryo fibroblasts (25%) could replace the feeder cell layer in recloning experiments with both human--mouse and mouse--mouse hybridomas. Therefore, conditioned media (prepared in large quantities) may be used for generating standardized conditions for high-efficient cloning and recloning of hybridoma cell lines.

Animals↗
PubMed1987

[Modification of in vitro immune mechanisms by various glucocorticosteroid preparations--studies of lymphocytes in healthy probands and autoimmune patients].

The influence of steroids on lymphocyte responses was tested in vitro. The inhibition of early events of lymphocyte stimulation and cell proliferation after PHA stimulation was estimated. We found, however, different individual responses of immune cells of various donors to the preparations dexamethasone, prednisolone, hydrocortisone or methylprednisolone in both in vitro models tested. Steroid treatment of lymphocyte cultures resulted in both inhibition and enhancement of PWM-driven Ig synthesis in cultures derived from different test persons. In experiments with lymphocytes from patients with acute systemic lupus erythematosus we found a dramatically enhanced PWM-stimulated IgG synthesis which is disturbed in cultures of these patients. The individuality of steroid sensitivity of lymphocyte cultures of healthy test persons and the abnormal reactivity of SLE lymphocytes were discussed in relation to therapeutical applications.

Adult↗
PubMed1987

The role of dipeptidyl peptidase IV in human T lymphocyte activation. Inhibitors and antibodies against dipeptidyl peptidase IV suppress lymphocyte proliferation and immunoglobulin synthesis in vitro.

Dipeptidyl peptidase IV (DP IV), an ectoenzyme in the cell membrane of T lymphocytes, is an important constituent in the process of lymphocyte activation. This conclusion is drawn from the following observations: (a) Proliferation of peripheral blood lymphocytes induced by mitogenic lectins (phytohemagglutinin, concanavalin A, pokeweed mitogen) is suppressed in the presence of DP IV inhibitors. This effect is specific and applies to an irreversible suicide inhibitor as well as to a competitive one in a dose-dependent fashion. (b) Inhibition of DNA synthesis was also induced by a polyclonal anti-DP IV immunoglobulin. (c) As a consequence of impaired T cell function the production of immunoglobulins by pokeweed mitogen-stimulated lymphocytes is also markedly reduced in the presence of DP IV inhibitors.

Antibody Formation↗
PubMed1987

The influence of interferon-gamma, interleukin-2, prostaglandin E2, and cyclosporine on the polyclonal and anti-DNA antibody secretion in lymphocyte cultures derived from patients with systemic lupus erythematosus.

The influence of various immunoregulatory substances was studied in lymphocyte cultures derived from patients suffering from systemic lupus erythematosus (SLE) by using the model of spontaneous secretion of polyclonal immunoglobulin G (IgG)/immunoglobulin M (IgM) and anti-DNA autoantibodies. Compared with healthy donors, lymphocytes derived from patients with active SLE disease showed an elevated secretion of total IgG as well as anti-DNA-IgG in vitro, which was associated with an increase in the proportion of activated (HLA-class II +) T cells in their peripheral blood. Recombinant interferon-gamma increases the total IgG/IgM as well as anti-DNA-IgG/IgM secretion, which suggests that it has a possible role in the pathogenesis of SLE disease. Recombinant interleukin-2 and prostaglandin E2 normalize the high, spontaneous total IgG secretion, but elevate anti-DNA-IgG/IgM secretion. These results suggest that autoreactive B-cell clones are regulated differently in SLE patients. Cyclosporine inhibits total IgG/IgM secretion in all patients and anti-DNA-IgG/IgM secretion in six of eight patients. The possible therapeutic use of such immunomodulatory substances in SLE disease is discussed.

Adolescent↗
PubMed1987

Cell biology of human IgM-producing hybridomas derived from a fusion of human spleen lymphocytes with mouse myeloma cells.

Hybrids were derived from the fusion of mouse myeloma cells with human spleen cells from a patient with active idiopathic thrombocytopenia. Of 288 initially seeded cultures, 186 were found to produce human Ig. The growth and Ig production rates, cloning efficiencies using different feeder layers and the karyotype were determined for 9 clones that stably produced human monoclonal IgM (2-100 micrograms/ml) for at least 9 months. All cells of the Ig-producing hybridoma clones were positive for cytoplasmic-Ig, whereas only 20-65% of cells expressed surface Ig (mu and chains). Human monoclonal antibodies in mass cultures were derived in serum-free PRMI 1640 medium. Two clones produced human IgM (nearly 2 mg/ml) in the ascitic fluid of nude mice. Feeder cells of peritoneal macrophages from Balb/c mice enabled more efficient recloning of human x mouse hybrids than did thymocytes. Nearly all subclones derived from 2 clones were found to produce the same monoclonal antibodies as the parental lines. Information on the individual parameters of a hybridoma cell line may be helpful in the large-scale production of human monoclonal antibodies.

Animals↗