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Biomedical subjects

J Wijdenes

Publications and source records attributed to J Wijdenes.

At least 163 records · Page 9Linked to original sources
PubMed1988

A monoclonal anti-IgE antibody against an epitope (amino acids 367-376) in the CH3 domain inhibits IgE binding to the low affinity IgE receptor (CD23).

We have produced three different mAb specific for human IgE-Fc. Their binding pattern to either heat-denatured IgE or a family of overlapping IgE-derived recombinant peptides and their ability to affect interaction of IgE with its low affinity receptor Fc epsilon R2/CD23 demonstrate that they recognize distinct epitopes on the IgE molecule. All three mAb were able to induce basophil degranulation as measured by the induction of histamine release. mAb 173 recognizes a thermolabile epitope in the CH4 domain. It does not affect the binding of IgE to Fc epsilon R2/CD23. mAb 272 recognizes a thermostable epitope that maps to a sequence of 36 amino acids (AA) spanning part of the CH2 and CH3 domain and it does not affect the binding of IgE to Fc epsilon R2/CD23. mAb 27 recognizes a thermolabile epitope located on a 10 AA stretch (AA 367-376) in the CH3 domain. This area contains one N-linked oligosaccharide (Asn-371), but the antibody is not directed against carbohydrate because it binds to Escherichia coli-derived IgE peptides. mAb 27 inhibits the binding of IgE to Fc epsilon R2/CD23 but is still capable of reacting with IgE already bound to Fc epsilon R2/CD23. These data suggest that upon binding to Fc epsilon R2/CD23, the IgE molecule engages one of two equivalent-binding sites close to the glycosylated area of the CH3 domain.

Animals↗
PubMed1988

Evidence for a polypeptide segment at the carboxyl terminus of recombinant human gamma interferon involved in expression of biological activity.

A panel of 18 murine monoclonal antibodies was raised in BALB/c mice to the full-length, 146 amino acid residue recombinant human gamma interferon (rHuIFN gamma-A). Two monoclonal antibodies, designated 47N3-6 and 30N47-1, were purified from ascites tumors and further characterized. Antibody 47N3-6 neutralized both the antiviral and antiproliferative activities of rHuIFN gamma-A. Both Western blotting and enzyme-linked immunosorbent assays indicated that antibody 47N3-6 could bind to rHuIFN gamma-A as well as to a genetically engineered truncated form lacking the first three amino-terminal residues (rHuIFN gamma-D) but did not recognize a genetically engineered variant terminating at residue 131 (rHuIFN gamma-B). This antibody also demonstrated binding to a 15 amino acid residue oligopeptide, designated F-1, corresponding to residues 132-146 at the carboxyl terminus of rHuIFN gamma-A. Chemical cleavage of peptide F-1 with cyanogen bromide produced two fragments that were separated by reversed-phase high-pressure liquid chromatography. Dot-blot analysis indicated that antibody 47N3-6 could bind to a fragment, KRKRSQHse, derived from residues 132-137 of rHuIFN gamma-A, but could bind only weakly to the cyanogen bromide fragment corresponding to residues 138-146. It was consistent with these results that antibody 47N3-6 demonstrated binding to a form lacking the five carboxyl-terminal amino acids (rHuIFN gamma-D') but did not bind to a synthetic polypeptide corresponding to residues 138-146. Peptide F-1 exhibited neither antiviral nor antiproliferative activity, and it did not antagonize the antiviral activity of rHuIFN gamma-A.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1987

Enumeration of CR1 complement receptors on erythrocytes using a new method for detecting low density cell surface antigens by flow cytometry.

A sensitive enhancing system was developed for detecting low density cell surface antigen by flow cytometry. The system termed the 'super avidin-biotin system' (SABS) uses biotinylated antibody, phycoerythrin-streptavidin (StreptA-PE), biotinylated goat anti-streptavidin antibody, and StreptA-PE. CR1 complement receptor antigenic sites were quantified on erythrocytes from healthy individuals and patients with antibodies against human immunodeficiency virus (HIV) using SABS and a conventional radioimmunoassay (RIA) with monoclonal anti-CR1 antibody. As little as 50 sites/cell were detected using either SABS or RIA. Accurate quantification of CR1 antigenic sites was achieved within the range of 100-1300 sites/cell. Similar results were obtained using either of the two methods. Intra-assay and day-to-day reproducibilities using SABS were 2% and 12% respectively, comparable to those of conventional RIA measurements. In addition to enumeration of CR1 on erythrocytes for clinical purposes, the use of SABS may probably be extended to a wide number of situations where a sensitive detection of low density cell surface antigen is needed.

Acquired Immunodeficiency Syndrome↗
PubMed1987

Production and characterization of a monoclonal antibody specific for the human lymphocyte low affinity receptor for IgE: CD 23 is a low affinity receptor for IgE.

In the present study a gamma 1 kappa monoclonal antibody, Mab 25, specific for the receptor for the Fc fragment of IgE on lymphocytes (Fc epsilon RL) was established. This antibody was generated after fusion of spleen cells from mice immunized with the EBV-transformed lymphoblastoid cell line RPMI 8866, which is known to express Fc epsilon RL at high density. Mab 25 inhibits strongly the binding of IgE to RPMI 8866 cells and to other Fc epsilon RL-positive EBV-transformed lymphoblastoid cell lines. A 50% inhibition of IgE binding was observed at a Mab 25 concentration of 10 ng/ml. The binding of IgE was also inhibited by Fab fragments of Mab 25, suggesting that the inhibition is not simply due to steric hindrance or to an eventual binding through its Fc portion. Mab 25 only binds to cell lines expressing Fc epsilon RL. Mab 25 immunoprecipitated a single polypeptide with an apparent m.w. of 42 Kd, pI 4.9. The membrane molecule bound to and eluted from a Mab 25 immunoabsorbent had the same apparent m.w. and pI as the Fc epsilon RL purified from an IgE immunoabsorbent. Additionally, when RPMI 8866 cell lysates were cleared with Mab 25, no Fc epsilon RL could be bound to or eluted from an IgE immunoabsorbent. Mab 25 was found to weakly bind to a minor proportion of blood (1 to 4%), tonsil (2 to 9%) and spleen (4 to 5%) mononuclear cells with a low intensity. By double fluorescence analysis, most of the Fc epsilon RL-positive cells were found to be CD 20-positive B lymphocytes. The staining pattern of Mab 25 and the biochemical characteristics of the antigen detected by Mab 25 were comparable to those of the CD 23 Mab. The four CD 23 Mab MHM 6, PL 13, HD 50, and Tü 1 were found to inhibit the binding of IgE. PL 13 was found to totally inhibit the binding of Mab 25 to RPMI 8866 cells, whereas Tü 1 and MHM 6 only partially inhibited Mab 25 binding. HD 50 was unable to block the binding of Mab 25. The finding that different CD 23/Fc epsilon RL-specific monoclonal antibodies recognizing distinct epitopes have in common the capacity of inhibiting the binding of IgE suggests that upon binding they induce a conformational alteration of the Fc epsilon RL resulting in a loss of the IgE binding capacity. In conclusion, our data demonstrate that the CD 23 antigen is a low affinity receptor for IgE on lymphocytes.

Antibodies, Monoclonal↗
PubMed1987

In the snail Helix aspersa the gonadotropic hormone-producing dorsal bodies are under inhibitory nervous control of putative growth hormone-producing neuroendocrine cells.

In many Stylommatophora, dorsal bodies (DB) consist of cell groups dispersed in the connective tissue surrounding the cerebral ganglia. It has previously been shown that in these animals the DB cells are under inhibitory nervous control. In the present tissue culture experiment on Helix aspersa it is shown that the axons innervating the DB cells originate from the peptidergic cerebral green cells (CeGC). It is argued that the observed morphological relationship between the CeGC and the DB--the CeGc produce a growth hormone, the DB a female gonadotropic hormone--reflects the antagonism between growth and reproduction which has often been reported for pulmonate snails.

Animals↗
PubMed1986

A flow cytometric micromethod for the detection of Fc epsilon receptors and IgE binding factors using fluorescent microspheres.

Two assays based on the use of fluorescent microspheres have been developed in order to detect Fc epsilon receptors on human cells and human IgE binding factors. A direct assay using microspheres to which IgE was coupled permitted the detection of Fc epsilon receptors on RPMI 8866 cells. However the fluorescence intensity was relatively low and made it difficult to discriminate between positive and negative cells. To increase the sensitivity of the assay, an indirect 3-step test was developed, in which the cells were subsequently incubated with soluble IgE, a polyclonal or monoclonal anti-IgE antibody and fluorescent microspheres to which anti-mouse or anti-rabbit immunoglobulin was coupled. This indirect assay resulted in an increased fluorescence intensity. Optimal discrimination between positive and negative cells was obtained. This assay permitted the detection of human IgE binding factors produced by RPMI 8866 cells. The binding of IgE was not dependent on the origin of the latter. Among the different cell lines tested, the EBV positive lymphoblastoid cells were found to express Fc epsilon receptors and to release IgE binding factors in their supernatants.

Antibodies, Anti-Idiotypic↗
PubMed1983

Effects of two gonadotropic hormones on polysaccharide synthesis in the albumen gland of Lymnaea stagnalis, studied with the organ culture technique.

A medium for culturing organs of the basommatophoran freshwater snail Lymnaea stagnalis is described. One of the accessory sex glands, the albumen gland, which produces proteins and galactogen as nutritive substances for the embryos, was cultured along with parts of the central nervous system (CNS), including the female gonadotropic hormone-producing dorsal bodies (DB). Culture time was 4 days. The cerebral ganglia and the DB appeared to have a strong stimulating influence on polysaccharide synthesis in the albumen gland. The remaining part of the CNS has a weak stimulating effect. The use of extracts (culture time 6-26 hr) of parts of the cerebral ganglia and of the DB showed that not only the dorsal body hormone (DBH), but also the ovulation hormone (CDCH)--a neurohormone produced by the caudodorsal cells located in the cerebral ganglia--stimulates the synthetic activity of the albumen gland. The action of these hormones is direct; i.e., it is not exerted via the gonad. Dose- and time-response relations for the DBH and the CDCH are very similar. No additional rise in stimulation occurred when the two hormones were administered together. Experiments with albumen glands of adult snails demonstrated that the presence of large quantities of secretory material in the gland inhibits the response to DBH and CDCH.

Animals↗
PubMed1983

Endocrine control of galactogen synthesis in the albumen gland of the slug, Limax maximus.

An in vitro method for culturing Limax maximus albumen glands is described in which the biosynthetic activity of the slug albumen gland was monitored by measuring the incorporation of [14C]glucose into galactogen. Homogenates of the central nervous system were shown to cause a 3.5- to 12-fold increase in galactogen synthesis in albumen gland explants as compared to controls. The major sources of the galactogen-synthesis stimulating factor (Gal-SF) were found to be the cerebral ganglia and their surrounding connective tissue. Gal-SF was demonstrated to be peptidase sensitive and heat labile suggesting that it is probably a polypeptide. Autoradiographs of semithin araldite sections supported the incorporation data: in albumen gland explants cultured with cerebral ganglion homogenate considerably more label was found over secretory granules than in control-cultured explants. The possible cellular source of Gal-SF is discussed in relation to its possible origin in other investigated pulmonates.

Albumins↗
PubMed1980

A comparative study on neurosecretion demonstrated by the alcian blue-alcian yellow technique in three terrestrial pulmonates (Stylommatophora).

Neurosecretion was studied in the central ganglia of three terrestrial pulmonates (Deroceras reticulatum, Arion hortensis, and Helix aspersa) with the alcian blue-alcian yellow (AB/AY) staining method. A number of neurosecretory cell types were distinguished in all three species (D. reticulatum: 9, A. hortensis: 11, H. aspersa: 13) With AB/AY the cells stain different shades of green and yellow. The histochemical differences reflect ultrastructural differences: the cell types contain elementary granules of different size and appearance. Some cell types form distinct groups of up to 50 cells, while others occur in smaller groups or as single cells. Neurosecretory axons could only be traced in a few cases to the probable neurohaemal areas. Based on the following criteria: (1) staining properties of cells (2) size of cells, (3) position of cells in the ganglia, it appears that the neurosecretory systems of the three species show great similarity, although some differences are apparent.

Animals↗
PubMed1980

The effect of thiol compounds on the autolysis of papain.

Incubation of papain with 3--17.5 mM dithiothreitol and dithioerythritol at pH 8.5 causes inactivation owing to autolysis. Such inactivation is not observed on incubation with equivalent concentrations of mercaptoethanol, cysteine or 2,3-dimercaptopropanol. The inactivation rate is independent of papain concentration within the range of 0.5--2% and is proportional to dithiothreitol concentration. This is in agreement with a sequence of two reactions: (Formula: see text) the first reaction being rate-limiting. S-Carboxymethyl-papain (I) and S-carboxamidomethyl-papain (II) were incubated with 18 mM dithiothreitol at pH 8.5 and, after stopping the reaction with iodoacetic acid, were subjected to gel electrophoresis. Electrophoretograms of both I and II exhibited a small new band attributable to a species with one disulphide bond reduced and carboxymethylated. The new band on II was more pronounced than that of I. It is argued that (Formula: see text) is a papain species with one reduced disulphide bond, sensitive to proteolytic attack by native papain.

Dithioerythritol↗
PubMed1979

A study of gonadal organogenesis, and the factors influencing regeneration following surgical castration in Deroceras reticulatum (Pulmonata:Limacidae).

At hatching, the hermaphrodite duct of Deroceras reticulatum consists of a single cell type designated the Gonadal Stem Cell (GSC). Proliferation of the GSC leads to the formation of numerous ductules each of which forms one of the acini of the gonad. The germinal and supporting cells are derived entirely from the GSC. The germ cells differentiate first, followed by the Sertoli and follicle cells. At the early sperm stage of gonadal development the hermaphrodite duct differentiates to function as a seminal vesicle. Once the GSC are committed to this change they lose their regenerative ability. The only remaining GSC are the cells of the acinar epithelium, and these retain their germinal potential until the death of the animal. Regeneration will occur from the hermaphrodite duct provided it is in the immature state, i.e., composed of GSC, and is exposed to the hormonal conditions of a young animal. Nervous connections and the presence of an artery are not necessary for this regeneration. The presence of a functional gonad does not inhibit regeneration.

Animals↗
PubMed1977

Dimerization of papain induced by mercuric chloride and a bifunctional organic mercurial.

The bifunctional mercurial meso-1,4-bis(acetatomercuri)-2,3-diethoxybutane and mercuric chloride are capable of dimerizing papain, by the attachment of the thiol group of two molecules of papain to each molecule of reagent. This is evident from the titration data, gel filtration and sedimentation equilibrium. The conformational change of papain necessary for this reaction is discussed.

4-Chloromercuribenzenesulfonate↗
PubMed1976

Proton equilibria in the binding of Zn2+ and of methylmercuric iodide to papain.

The proton liberation on the binding of zinc chloride and methylmercuric iodide to the (essential) thiol group of papain has been examined as a function of pH. This was carried out by (a) direct titration of the protons on the addition of the metal compound to active papain and (b) measurement of the extent of inhibition of enzyme activity by the metal compound as a function of pH. It was found that in the neutral pH range the thiol group or the neighbouring imidazole group in the free enzyme carries one proton, at low pH both groups do so, whereas at high pH neither group carries a proton. The pK values of the free enzyme that govern the proton release, 4.2 and 8.5, correspond to those that govern overall activity. Both from the experiments with methylmercuric iodide and from fluorescence measurements of methylmercuric papain, it was established that the imidazole group in the latter compound exhibits a pK of 5.4. Taking recent data into account, it was considered that the ion pair of thiolate anion and imidazolium cation, proposed by Polgar, is the best approximation to describe the charge distribution in the active centre and to explain the reaction mechanism.

Binding Sites↗
PubMed1976

An ultrastructural in vitro study on the regulation of neurosecretory activity in the freshwater snail Lymnaea stagnalis (L.) with particular reference to caudo-dorsal cells.

The neurosecretory Caudo-Dorsal Cells (CDC) in the cerebral ganglia of the freshwater pulmonate snail Lymnaea stagnalis produce an ovulation stimulating hormone. Previously it has been shown that neuronal and non-neuronal inputs are involved in the regulation of their activity. The degree of autonomy of these cells has been investigated by studying with morphometric methods the ultrastructure of CDC maintained in vitro. CDC of isolated cerebral ganglia which were cultured for 7 days show a considerable rate of synthesis, transport and release of neurohormone. Apparently these processes can proceed in the absence of neuronal and hormonal inputs from outside the cerebral ganglia. Completely isolated CDC, however, do not show neurosecretory activity in vitro; active Golgi zones, indicating the formation of neurosecretory elementary granules, are absent from such cells. Isolation does not seem to affect general cell functions such as protein synthesis and respiration. It is suggested that a neuronal input, originating within the cerebral ganglia, is necessary for the stimulation of CDC neurosecretory activity. Techniques are described for the isolation and culture of neurosecretory cells of L. stagnalis.

Animals↗