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Biomedical subjects

A H Sehon

Publications and source records attributed to A H Sehon.

At least 55 records · Page 3Linked to original sources
PubMed1987

Tolerance induction in mice by conjugates of monoclonal immunoglobulins and monomethoxypolyethylene glycol. Transfer of tolerance by T cells and by T cell extracts.

The specific tolerance induced in mice by conjugates of human monoclonal IgG (HIgG) with monomethoxypolyethylene glycol (mPEG) was transferred to normal mice by spleen cells or a surface immunoglobulin negative (sIg-) Lyt-2+ subpopulation of these cells. Although transferable tolerance was demonstrable 6 to 14 days after treatment of the cell donors with tolerogen, the state of tolerance persisted in the treated mice for at least 43 days. Moreover, an extract prepared by freezing and thawing of the sIg- spleen cells obtained from mice 6 days after treatment with HIgG(mPEG)20 was capable of reducing (greater than 85%) the immune response of normal mice to heat aggregated HIgG. On the basis of these results, it is suggested that similar tolerogenic mPEG derivatives of xenogeneic monoclonal immunoglobulins (XIg) may prove to be useful therapeutic agents in man when administered before treatment with the unmodified XIg.

Animals↗
PubMed1987

Recognition of a site of a Kentucky bluegrass pollen allergen by antibodies in the sera of allergic and non-atopic humans and a murine monoclonal antibody.

Allergen 27 was isolated from the aqueous extract of Kentucky Bluegrass pollen (KBG-R) with a reversed immunosorbent prepared by coupling murine monoclonal antibody, Mab 27, to Sepharose 4B. Sera of patients allergic to KBG pollen, as well as serum of nonatopic individuals possessing anti-KBG antibodies, inhibited the binding of Mab 27 to either Ag 27 or KBG-R to the extent of 20 to 35% in ELISA. In contrast, sera devoid of antibodies to KBG-R had no inhibitory capacity. In a radioallergosorbent test, it was demonstrated that Mab 27 could inhibit the binding of human IgE antibodies to Ag 27 to the extent of 52%. From these results, it is concluded that Ag 27 contains a determinant recognized by both human IgE and blocking antibodies and a murine Mab.

Allergens↗
PubMed1987

Induction of in vivo helper activity for murine antibody responses by macrophages pulsed with ovalbumin-monomethoxypolyethylene glycol (OA-mPEG) conjugates.

Conjugates of protein antigens with an optimal number of monomethoxypolyethylene glycol (mPEG) chains of an appropriate molecular weight had been shown to suppress murine IgE responses to the unmodified antigen. To investigate the possibility that the tolerogenic capacity of these mPEG conjugates is attributable to a defect in macrophage (M phi) presentation of their antigenic determinants, the activity of ovalbumin (OA)-mPEG conjugates when pulsed onto mouse peritoneal adherent cells (M phi) was compared in this study with their activity in solution. Surprisingly, in contrast to the suppressogenic capacity of mPEG conjugates in solution, the OA-mPEG pulsed M phi appeared to exert a helper effect when injected intraperitoneally (ip), i.e., after subsequent immunization with dinitrophenylated OA (DNP3-OA) in Al(OH)3, the mice showed accelerated IgE and IgG1 antibody responses to OA and DNP. However, when M phi were exposed to limiting concentrations of OA or OA-mPEG, markedly higher concentrations of OA-mPEG were required to yield pulsed M phi, exerting a significant helper effect. It was concluded that although M phi were capable of presenting the OA determinants of OA-mPEG conjugates to helper T (Th) cells, the preparations of modified antigen were presented less effectively than native OA.

Animals↗
PubMed1987

Tolerogenic polyethylene glycol derivatives of xenogeneic monoclonal immunoglobulins.

It is becoming apparent that the effectiveness of xenogeneic monoclonal antibodies (XIg), which are increasingly used for diverse therapeutic purposes in man, may be counteracted by their inherent immunogenicity. Since conjugates of proteins with monomethoxypolyethylene glycol (mPEG) have proved to be effective tolerogens in other systems, we have used an experimental model in mice to explore the tolerogenicity of mPEG conjugates of a human monoclonal IgG (HIgG), i.e. a myeloma protein. Administration of these conjugates prior to immunization with heat aggregated HIgG (ha-HIgG) resulted in specific tolerance, as manifested by a marked reduction in the level of antibodies to HIgG, which was related to the degree of conjugation and the dose of conjugate administered. Thus, administration of HIgG(mPEG)20 6 to 43 days prior to immunization with ha-HIgG resulted in an inhibition of anti-HIgG antibody formation of the order of 85-90%, in relation to the titres of mice receiving PBS in lieu of HIgG(mPEG)20; these results hold the promise that mPEG conjugates of XIg may prove therapeutically useful in man in relation to organ transplantation, localization of tumours by immuno-imaging and tumour destruction by immunotoxins.

Animals↗
PubMed1987

Production and characterization of human-human and human-mouse hybridomas secreting Rh(D)-specific monoclonal antibodies.

Human monoclonal antibodies specific for the Rh(D) antigen were produced by cell lines generated by the fusion of pooled Epstein-Barr virus (EBV)-transformed B-cell lines secreting Rh(D) antibodies with the murine myeloma cell line NS.1 or with the human lymphoblastoid cell line HOA.1. The selection of hybrids was achieved in RPMI 1640 medium containing HAT and ouabain. Higher fusion efficiency was obtained with the NS.1 cell line; however, the hybrids with HOA.1 exhibited a greater clonal stability. The products of four clones (three human-human and one human-mouse) that consistently secreted antibodies for over 11 months were tested for specificity with a panel of red cells of various Rh phenotypes. The supernatants of all four clones showed anti-Rh(D) specificity but failed to react with the red cell Du phenotypes categorized as DV(Dw+) and DVI. Two of the three human-human clones secreted IgM(lambda) and the third IgG(kappa). The human-mouse clone produced IgG(kappa) antibody.

Animals↗
PubMed1987

A canine model for the study of hapten-specific suppression of IgE-mediated bronchoconstriction and anaphylaxis.

Newborn mongrel dogs were sensitized with conjugates of ovalbumin (OA) and 2,4-dinitrophenol (OA-DNP3) in the presence of Al(OH)3 to produce high levels of anti-OA and anti-DNP IgE antibody. At 4-6 months of age, when anti-DNP and anti-OA antibody levels reached titers of 64 by passive cutaneous anaphylaxis, the dogs underwent separate inhalation and intravenous challenges with conjugates of DNP and bovine gamma globulin (DNP15-BGG) and OA. Inhalation challenge with DNP15-BGG and OA resulted in 5- and 10-fold increases in airflow resistance, respectively. Intravenous challenge with either DNP15-BGG or OA produced profound anaphylaxis with 60-80% decreases in blood pressure, cardiac output and regional blood flows in the carotid, superior mesenteric and renal arteries, and the distal aorta. Treatment of sensitized dogs with 5 doses of 20 mg of conjugates of DNP and polyvinyl alcohol (DNP2-PVA) on alternate days resulted in suppression of anti-DNP IgE antibody production; abrogation of established airway and vascular anaphylactic sensitivities; no change in regional blood flows, and no effect on sensitivities to challenge with OA.

2,4-Dinitrophenol↗
PubMed1986

Production and characterization of a monoclonal antibody to biotin.

Monoclonal antibodies to biotin have been prepared by using biotin linked to keyhole limpet haemocyanin (KLH) as the antigen. Spleen cells obtained from mice immunized with biotin-KLH were fused with the myeloma cell line NS-1. The resulting hybridomas were screened for the production of antibodies to biotin using an enzyme-linked immunosorbent assay. Clones producing antibodies to biotin were isolated by limiting dilution methods. Four cell lines, each derived originally from a different fusion, were chosen for the production of monoclonal antibodies. The monoclonal antibodies obtained have been characterized with respect to their ability to interact with biotin, biotin-bovine serum albumin, biotin-KLH and biocytin as well as to inhibit biotin-dependent enzymes. They have been used to produce cellular biotin deficiency in vitro for studies of biotin function.

Animals↗
PubMed1986

The influence of antibody affinity on the radioallergosorbent test (RAST) and in vitro histamine release. Studies with hapten-specific monoclonal IgE antibodies.

Four murine monoclonal IgE antibodies specific for the hapten, 4-hydroxy-3-nitrophenylacetyl (NP), had been previously found to be heteroclitic in nature in that they bound the crossreacting hapten, 4-hydroxy-3-iodo-5-nitrophenylacetyl (NIP), with greater affinity than NP. The influence of antibody affinity on the results of two commonly used assays for IgE, namely the radioallergosorbent test (RAST) and histamine release from rat peritoneal mast cells, was studied using these antibodies. In general, in agreement with previous reports, it was found that affinity influences both RAST and histamine release; however, the affinity constants deduced from equilibrium dialysis measurements for the reactions with monovalent haptens were not directly related to the activities of the antibodies as reflected in assays using multivalent hapten protein conjugates.

Animals↗
PubMed1986

Partial characterization of an antigenic site of high molecular weight basic antigen, a ryegrass pollen allergen, using a monoclonal antibody.

We have previously reported on the production of murine monoclonal antibodies (Mabs) to the retentate fraction of the aqueous extract of Kentucky bluegrass pollen (KBG-R) [Kisil et al. (1980) Fedn Proc. 39, 3479]. In the present study, the ability of one of these Mabs (Mab 12) to recognize various antigenic components present in KBG-R and the corresponding fraction of ryegrass (rye-R) was evaluated by the Western and immunoblot methods. Thus, KBG-R and rye-R were resolved electrophoretically into a large number of components. Remarkably, the concurrent immunoblot analysis with Mab 12 detected only a single antigenic component in each of the retentate fractions. The position of the antigenic component observed on these immunoblots was identical to that obtained with the rye allergen high mol. wt basic antigen (mol. wt 56,800). To characterize the antigenic site recognized by the Mab, the size of HMBA was reduced by cleavage with CNBr, the resulting fragments separated by high-performance liquid chromatography on a reverse-phase column and their antigenic activity analyzed by immunoblot. Two peptides, CB-1 (mol. wt = 17,400) and CB-2 (mol. wt = 22,000), retained the capacity to react with Mab 12 and also IgE antibodies present in a pool of sera from grass allergic individuals.

Antibodies, Monoclonal↗
PubMed1986

Proliferative responses by immune murine popliteal lymph node cells to high-molecular-weight basic allergen of ryegrass pollen and its cyanogen bromide fragments.

High-molecular-weight basic allergen (HMBA) of ryegrass pollen and its fragmented form obtained by cleavage with cyanogen bromide (CNBr) (HMBA-frag) were evaluated for their immunogenicity in terms of their capacity to stimulate the in vitro proliferative responses of the in vivo antigen-primed popliteal lymph node (PLN) cells. For this purpose, mice (C3H/HeJ) were immunized in the hind foot pads with solutions containing 1-5 micrograms of HMBA or HMBA-frag emulsified in Freund's complete adjuvant (FCA). Eleven days later, single-cell suspensions of the PLN were cultured in vitro for 4 days in the presence of either HMBA or HMBA-frag at concentrations ranging from 0.625 to 20 micrograms/ml. 3H-thymidine was added to the cultures 6 h prior to harvesting the cells. The degree of proliferation was assessed from the extent of intracellular incorporation of the 3H-thymidine by the PLN cells. Comparable degrees of lymphocyte proliferation were consistently obtained on stimulation with either HMBA or HMBA-frag of PLN cells from mice immunized with the allergen HMBA. A similar finding was made using PLN cells from mice immunized with the HMBA-frag. This study demonstrated that the CNBr-derived fragments of HMBA retained the immunogenicity of the parent molecule HMBA in terms of being able to (i) induce the in vivo priming of PLN cells and (ii) elicit the in vitro proliferative response of the antigen-primed PLN cells.

Allergens↗
PubMed1986

Isolation of a Kentucky Blue grass pollen allergen using a murine monoclonal antibody immunosorbent.

We have previously reported the production of murine monoclonal antibodies to the retentate fraction of Kentucky Blue Grass pollen (KBG-R). In the present study, one of these monoclonal antibodies (Mab 27) was used in a combination of methods employing Western and immunoblot analysis to establish its reactivity to various antigenic components present in KBG-R. Thus, Mab 27 reacted predominantly with an antigen having a MW of 30 kD and to a lesser extent with other antigenic components with MW of 35 and 17 kD. Fractions of KBG-R obtained by preparative isoelectrofocusing, revealed on analysis by ELISA, that Mab 27 reacted with several components differing in charge. From these observations it was concluded that KBG-R contained a group of related antigens detectable with Mab 27 and differing in size and charge. A reverse immunosorbent, prepared by coupling Mab 27 to CNBr-activated Sepharose 4B, was used to absorb KBG-R. Bound material was recovered by acid elution and designated as Ag 27. SDS-PAGE analysis revealed that Ag 27 consisted of at least two components with molecular weights of 30 and 17 kD. Analysis by RAST using sera from humans allergic to KBG indicated that Ag 27 was highly allergenic.

Allergens↗
PubMed1985

Demonstration of intracytoplasmic IgE in circulating lymphocytes of allergic individuals.

Cells containing intracytoplasmic IgE (IC IgE) were demonstrated in the peripheral blood mononuclear cells of atopic individuals by means of indirect immunofluorescence, employing mouse monoclonal anti-human IgE antibody. IC IgE-positive cells are low-density B cells as shown by centrifugation on discontinuous bovine serum albumin density gradient and by their reactivity with B1 monoclonal antibody specific to B lymphocytes, respectively. Further characterization of these cells by means of a rosette assay employing anti-Ig-coupled bovine erythrocytes indicated that these cells were surface IgE (sIgE) positive and sIgM negative. The data strongly suggest that activated IgE B cells are circulating in the blood of allergic individuals.

B-Lymphocytes↗
PubMed1985

Induction of IgA-specific suppressor activity in human T-lymphocyte cultures.

Cell-free supernatants of human circulating T-lymphocyte cultures incubated with secretory IgA (S-IgA) specifically suppressed both spontaneous IgA synthesis by B lymphocytes isolated from allergic individuals and pokeweek mitogen-induced IgA secretion by peripheral blood mononuclear cells. Cell-free supernatants of T-cell cultures incubated with IgE had no effect on IgA, IgG, or IgM synthesis. Hence, it is concluded that upon incubation with S-IgA, but not with another Ig class, T lymphocytes release IgA-specific suppressor factors.

Antibody Specificity↗
PubMed1985

Human cord blood suppressor T lymphocytes: I. Phenotype and target of the inducer of suppressor cell factor.

The present study examines the characteristics of the cord suppressor T cells and their targets. When an inducer of suppressor T cell factor (TisF) from culture supernatant of alloantigen-activated cord lymphocytes was used as a source of suppression, adult lymphocyte response to soluble antigen candidin, alloantigens, and pokeweed mitogen were all suppressed. These results suggest that TisF acts primarily on T cells in an antigen-nonspecific fashion. Furthermore, preculture T cell irradiation, but not T8+ cell depletion, of adult lymphocytes abrogated the suppressive effect of TisF on the PWM-stimulated IgG production, suggesting that the target of TisF belongs to the radiosensitive T8-(T4+) T cell subset. When different cord T cell subsets were tested for suppressor activity, only radiosensitive T4+ cells were capable of suppressing IgG production by adult lymphocytes. Neither preculture T8+ cell depletion nor irradiation of adult T cells affected the suppression. Taken together, these results suggest that the suppressor activity of cord T cells originated from an inducer of suppressor cells that activates radiosensitive T4+ suppressor effector cells through TisF. In turn, these effector cells act on either radioresistant T4+ helper T cells or B cells or both.

Adult↗
PubMed1985

Immunomodulation in rats by transplantable anterior pituitary tumors.

The effect on the immune response of the MtT/W5 (W5), MtT/W10 (W10) tumors of Wistar-Furth rats and of MtT/F4 (F4) tumor transplantable in Fischer 344 rats was examined. Antibody response to sheep red blood cells and contact sensitivity reactions to dinitrochlorobenzene (DNCB) were used as immune parameters. In intact rats the W5 tumor suppressed the antibody response as much as did hypophysectomy (Hypox). The antibody response of Hypox rats was marginally improved by this tumor. Contact sensitivity was not suppressed in intact animals and the poor reactivity of Hypox rats was reconstituted to normal levels by W5. Treatment with bromocriptine (BRC) had no influence on tumor growth or on the immune reactivity of tumor-bearing hosts. The W10 tumor suppressed antibody formation, but not contact sensitivity in intact animals; the antibody and the DNCB response of Hypox animals was reconstituted partially, by this tumor. BRC treatment of tumor-bearing animals mimicked the effect of Hypox in this system. The F4 tumor suppressed antibody formation in intact rats and did not reconstitute the reactivity of Hypox rats. The DNCB response was not influenced by this tumor in intact animals and it was partially reconstituted in Hypox rats. Again, the reactivity of BRC-treated animals was similar to that of Hypox tumor-bearing animals. These results indicate that pituitary tumors may alter the immune reactivity of their hosts significantly.

Adenoma↗
PubMed1985

Detection and characterization of monoclonal antibodies specific to IgE receptors on human lymphocytes by flow cytometry.

BALB/c mice were immunized with human lymphoblastoid cells (RPMI 8866 cells) expressing surface receptors for IgE (Fc epsilon R). Spleen cells from animals displaying high titres of anti-Fc epsilon R antibodies were fused with HGPRT-deficient NSI myeloma cells. Anti-Fc epsilon R antibodies were identified by a flow cytometric assay based on their ability to block the binding of IgE-coated fluorescent latex particles to Fc epsilon R-positive cells. Fourteen monoclonal hybridoma cell lines secreting antibody of the required specificity were amplified in tissue culture and then grown in the peritoneal cavity of BALB/c mice in order to obtain ascitic fluids with high antibody titres. The specificity of each monoclonal antibody (Mab) to lymphocyte Fc epsilon R was shown by the following observations: (i) the intact monoclonal antibody molecule or, in some cases, its F(ab')2 fragments blocked the binding of IgE to several Fc epsilon R(+) cell lines different from that employed for the initial immunization; (ii) the Mab bound directly to all the Fc epsilon R(+) cell lines tested, but not to several Fc epsilon R(-) cells as determined by indirect immunofluorescence; (iii) the binding of Mab to Fc epsilon R(+) cells was selectively blocked by IgE, but not by the other classes of Ig; and (iv) Mab had no effect on the binding of IgG to Fc gamma R on normal human peripheral blood mononuclear cells (PBMC).

Animals↗