Biomedical subjects

N R Ling

Publications and source records attributed to N R Ling.

At least 37 records · Page 2Linked to original sources

PubMed1987

ELISA measurement of IgG subclass production in culture supernatants using monoclonal antibodies.

Specific and sensitive ELISA to quantitate the human IgG subclasses in cell culture supernatants are described. These assays detect a minimum of 5 ng/ml IgG1, 90 ng/ml IgG2, 8 ng/ml IgG3 and 8 ng/ml IgG4 and can generally measure IgG subclasses in lymphocyte cultures containing a minimum of 200 ng/ml of total IgG. The isotype specificity of these ELISA is demonstrated and each individual ELISA shown to react with a number of paraproteins of the relevant subclass independently of their light chain type or their (major Caucasian) allotype. These assays have been used to determine the IgG subclass response of normal human lymphocytes to pokeweed mitogen in vitro.

Antibodies, Monoclonal↗

PubMed1987

Human neoplastic B cells express more than two isotypes of immunoglobulins without deletion of heavy-chain constant-region genes.

Flow cytometric analyses of surface immunoglobulins of human neoplastic B cells from four patients indicated that more than two isotypes were expressed on the surface of each patient's neoplastic B cells. Southern blot analysis of DNAs of these cells showed that each patient's neoplastic cells had the monoclonal rearrangement profile of the J segment of the immunoglobulin heavy-chain gene. Heavy-chain constant-region genes of these cells had no deletion associated with S-S recombination. Northern blot analysis of RNA from two neoplastic cells revealed that each RNA contained comparable amounts of mRNAs for two different isotypes which were identified by surface staining. These results support the hypothesis that the simultaneous expression of two different isotypes in a certain stage of B-cell differentiation is mediated by alternative RNA splicing without DNA deletion.

B-Lymphocytes↗

PubMed1987

Modulation of the murine immune response to human IgG by complexing with monoclonal antibodies. I. Antibody responses to determinants on the constant region of light chains and gamma chains.

Complexing a human IgG lambda paraprotein with a monoclonal antibody (McAb) to a lambda-chain determinant markedly depressed the anti-lambda response of immunized mice. No anti-lambda was found in the serum after two or four injections of the complex, whereas high titres of anti-lambda chain antibody were found in the sera of mice immunized with the IgG paraprotein complexed with any single anti-gamma chain McAb or with a pool of anti-gamma chain McAbs. Responses to the highly immunogenic Fc gamma portion of the molecule were not suppressed by complexing with a single anti-gamma chain McAb and were only slightly suppressed after complexing with a pool of anti-gamma chain McAbs. Some anti-Fc gamma antibody was produced by all animals receiving a single injection of the immunogen complexed to any McAb, but free immunogen did not generate a primary response. Complexing the IgG with an anti-Fc gamma McAb enhanced the response to to the poorly immunogenic C gamma 1 domain but no anti-C-gamma 1 was produced by mice receiving the IgG complexed with an anti-C-gamma 1 McAb. In immunizations with 'free' (Bence-Jones) lambda chain, all the antibody produced was directed against 'free'-specific determinants. Complexing the Bence-Jones protein with McAbs to 'free' or 'general' determinants on the lambda chain did not enhance or suppress the response. It is concluded that the response to weakly or moderately immunogenic, but not to strongly immunogenic, regions of the molecule may be suppressed by complexing the immunogen with a McAb of appropriate specificity. Possible reasons for this result are discussed.

Animals↗

PubMed1987

Modulation of the murine immune response to human IgG by complexing with monoclonal antibodies. II. Antibody responses to idiotopes of the human IgG paraprotein and of the mouse monoclonal antibodies.

Anti-idiotope antibodies produced by mice immunized with a human IgG paraprotein complexed with various mouse monoclonal antibodies (McAbs) have been measured. All animals receiving more than one injection of the paraprotein (free or complexed with a mouse McAb) produced antibodies to the idiotypes of the paraprotein. Complexing with a McAb, especially an anti-Fc-gamma McAb, enhanced the response. Antibodies to the idiotopes of mouse McAbs were more difficult to produce and their production was very dependent on the mode and schedule of the immunization. The best antisera were produced by mice receiving a course of injections of pre-formed complexes of the IgG paraprotein and McAbs. Four of five mice produced antibodies to the idiotopes of an anti-light chain McAb (C4) after a course of immunization (one primary plus four boosts) of an IgG-C4 complex. Two of the six mice receiving a similar course of injections of the paraprotein complexed with an anti-gamma McAb (A55) produced high titres of antibodies to A55 idiotypes. Responses were enhanced when complexes were prepared with a pool of McAbs. It is probable that the formation of large multi-cross-linked complexes containing the McAb under study is important in generating the response. Once a response is initiated, very high titres may be achieved.

Animals↗

PubMed1986

Studies on the reappearance of MHC class II antigens on cells of a variant human lymphoblastoid line.

EB4 lymphoblastoid cell line cells were cultured for 32 days with a ricin A chain-conjugated monoclonal antibody (McAb) to a beta chain determinant present on DP and DR antigens. A single colony of cells which survived this treatment was grown up and surface expression of MHC class II and B cell antigens measured. All class II antigens (DQ as well as DP and DR, alpha as well as beta chains) were initially greatly diminished, but substantial recovery of expression occurred within 20-100 days of culture (approx. 21-105 generations), although recovery was still incomplete. The CD19 (p95) B cell antigen was present in greater amount and the CD22 B cell antigen and surface immunoglobulin in lower amount on the variant line cells. It was confirmed that unconjugated anti-class II McAb binds to surface antigens but is unable to induce modulation even over an 8-day culture period. Evidence is presented that the gradual re-expression of class II antigens on the variant line cells is not due to the appearance of a mutant or to recovery from modulation. It is suggested that the variant line cells produce an excess of a regulatory molecule when grown in the conjugate.

Antibodies, Monoclonal↗

PubMed1986

Use of monoclonal antibodies reactive with secretory epithelial cells for the immunocytochemical identification of plasma cells.

Six monoclonal antibodies (McAbs) were identified as plasma cell-reactive when screened on sections of human tonsil. They were all produced following immunisation of mice with cells of a human plasmacytoid line. Three of the antibodies also stained the cytoplasm (but not the surface) of blood B cells and were unreactive with other leucocytes; one McAb showed broad lymphocyte reactivity and two were completely unreactive with blood leucocytes; on testing with a panel of cell lines specificity for the plasmacytoid line was demonstrated by three of the McAbs. In spite of the marked restriction shown by the reactivity of these antibodies in tests on cells of haemopoietic origin, tests on other human tissues - including thyroid and pancreas - showed that a related antigen was present in the cytoplasm of secretory epithelial cells. The overall patterns of reactivity of the individual McAbs on various tissues and blood lymphocytes were different. Comparisons were made with the established McAb OKT10, which binds to plasma cells, early stem cells and activated lymphocytes; its binding to plasma cells was confirmed and it was shown that it did not stain secretory epithelia. The potent reactions obtained with the new McAbs suggest that antibodies to antigens associated with epithelial cell secretory apparatus provide potentially useful reagents for studying plasma cells.

Animals↗

PubMed1986

Autoantibodies to acetylcholine receptor in myasthenia gravis: light chains.

We studied the light chain type of autoantibodies to acetylcholine receptor (AChR) by affinity chromatography with monoclonal anti-kappa and anti-lambda antibodies. The autoantibodies in four of eight myasthenic patients were of a single light chain type; the others comprised both types. In Graves' disease and cold-reactive hemolytic anemia, the pathogenic autoantibodies are confined to a single light chain type in individual patients, and in other diseases, doubtfully pathogenic autoantibodies are invariably mixtures of both light chain types. AChR antibodies may comprise both pathogenic and nonpathogenic types of autoantibody.

Aged↗

PubMed1986

Immunogenic and antigenic epitopes of immunoglobulins. XVIII. Subpopulations of human lambda chains defined with a panel of monoclonal antibodies.

Following immunization with various lambda Bence-Jones proteins, over 100 hybridomas were established. A majority of the monoclonal antibodies (McAb) produced were demonstrated to recognize epitopes common to all lambda chains or unique to the immunogen. Twenty-nine McAb exhibited profiles of 'restricted' reactivity. Analysis with a panel of serologically defined antigens allowed McAb recognizing the lambda II subgroup to be identified. Other McAb recognized epitopes selectively expressed by lambda II and lambda III subgroup proteins. A further twenty-one McAb exhibited individual specificity profiles, some of which detect minor lambda chain subpopulations. Several of these McAb have potential for analysis of the clonality of specific antibody responses and for detecting monoclonal B-cell proliferations.

Antibodies, Monoclonal↗

PubMed1986

An antigenic study of human plasma cells in normal tissue and in myeloma: identification of a novel plasma cell associated antigen.

A mouse monoclonal antibody named BU11 which detects an antigen strongly expressed on human plasma cells is described. The antibody stains plasma cells in tonsil sections, fresh and cultured plasmacytoid cells from the bone marrow of patients with multiple myeloma and cells of the plasmacytoid cell line RPMI 8226 used as the immunogen. In vitro studies of pokeweed mitogen (PWM) stimulated peripheral blood B cells and Epstein-Barr virus (EBV) stimulated tonsil B cells show that the antigen is present mainly on cells coexpressing the OKT10 antigen and containing cytoplasmic immunoglobulin (cIg). The BU11 antigen is expressed weakly on some normal B cells and is not present on T cells, monocytes or granulocytes. The antigen is of molecular weight 58kD under reducing conditions and is biochemically distinct from previously described plasma cell antigens.

Animals↗

PubMed1986

Human follicular dendritic cells (FDC): a study with monoclonal antibodies (MoAb).

A collection of new and established FDC-reactive MoAb has been used in an immunohistological study designed to throw light on (a) the nature of FDC (which are of unknown lineage) as judged by their sharing of antigens with other cell types and (b) the reason for the strong expression of some B cell antigens on FDC. The MoAb were tested on: (1) sections of tonsil, (2) sections of lymph nodes from four cases of non-Hodgkin's lymphoma, (3) peripheral blood cells and (4) cells of cultured haemopoietic cell lines. Only one of the ten new MoAb bound to FDC and no other component of the tissues screened. It resembled R4/23, a MoAb known to be specific for FDC. The other nine antibodies showed a range of cross-reactivity patterns involving one or more of the following: monocytes, macrophages, platelets, epithelium, endothelium and connective tissue fibres. Some of the MoAb reacted with B lymphocytes and cells of B lymphoblastoid lines but none showed the restricted FDC-staining pattern associated with MoAb which detect the CD23, P45 antigen. The findings are discussed in terms of the intrinsic or extrinsic nature of the antigens detected.

Antibodies, Monoclonal↗

PubMed1985

Variants of lymphoid lines produced with ricin A-chain monoclonal antibody conjugates.

Conjugates of ricin A-chain with monoclonal anti-light chain antibodies specifically killed cells hearing kappa or lambda immunoglobulin (Ig) light chains. Exposure of cells from B-lymphoblastoid cell lines (B-LCL) to conjugate for less than 30 h had only a slight effect on cell growth, but on 48 h exposure a marked killing effect was achieved. After recovery of growth, cells were re-exposed to conjugate for 9-14 days. Treatment of cells from the EB4 line (sIgG lambda) in this way yielded 4 variants which showed a marked reduction in levels of surface Ig lambda and secreted Ig lambda with slight, or no, reduction in MHC class II expression and similar growth rates to the parent line. Variant lines retained their phenotype over long periods of culture.

Antibodies, Monoclonal↗

PubMed1985

Evaluation of monoclonal antibodies having specificity for human IgG sub-classes: results of an IUIS/WHO collaborative study.

Seventy-four monoclonal antibodies (McAb) of putative specificity for human IgG (11), the IgG sub-classes (59) or Gm allotypes (4) have been evaluated for reactivity and specificity in eight laboratories employing different assay techniques or protocols. For the IgG, IgG3, IgG4, G1m(f) and G3m(u) specificities McAb have been produced that can be satisfactorily applied in most methodologies employed and have potential as reference reagents. The IgG1 and particularly IgG2 specificities proved problematical with all McAb evaluated demonstrating apparent assay restriction and whilst performing well in some assays proved to be poor or inactive reagents in others. However, the study identifies McAb individually suited to application within most commonly employed methodologies. Epitope display is the probable variability rather than capricious behaviour by the McAb. IgG1 and IgG2 were the least immunogenic of the sub-class proteins and there is evidence that epitope display is influenced by the physical and chemical procedures used to immobilize or fix antigen - a common requirement in the assay systems studied.

Antibodies, Monoclonal↗

PubMed1985

Immunogenicity and antigenicity of immunoglobulins. XII. Intact light chain and heavy chain isotype-restricted Vk-associated epitopes.

Immunization with intact IgG has allowed the isolation of four hybridomas producing antibodies recognizing epitopes expressed within subpopulations of human kappa light chains unrelated to known polymorphisms (Km) and previously defined V-region subgroups. The V-region-associated epitopes recognized are conformation-dependent, being expressed on intact light chain but not on isolated VK or CK fragments. The frequency of expression within paraprotein panels of different heavy chain isotypes varied between individual antibodies. An epitope recognized by B2A6, expressed by greater than 85% IgGK paraproteins, was not represented in 16 IgM paraproteins tested, suggesting that association of VK with mu chains does not result in display of the epitope recognized, or alternatively, that selective association between VK and CH gene products occurs. These data contrast with the reactivity of other McAb for CK epitopes which were reactive with isolated CK fragments, and for all kappa-bearing paraproteins, regardless of heavy chain isotypes.

Antibodies, Monoclonal↗

PubMed1984

The separation of human serum IgG into subclass fractions by immunoaffinity chromatography and assessment of specific antibody activity.

Murine monoclonal antibodies ( McAbs ) with specificity for subclass-specific or subclass-restricted determinants on human IgG have been coupled to Sepharose to generate affinity columns. The judicial use of positive and negative chromatography and the exploitation of the special properties of individual McAb affinity columns has allowed the preparation of individual IgG subclasses from polyclonal IgG containing less than 1% contamination by any other IgG subclass. The specificity of the antibodies present in each polyclonal IgG subclass preparation has been assayed against a bacterial toxoid (tetanus), 2 bacterial cell wall antigens (E. coli and pneumococcal) and coat antigen(s) of a DNA virus (CMV). Antibodies were predominantly IgG1 to tetanus toxoid, IgG2 to pneumovax and E. coli cell walls, and IgG1, 2 and 3 to CMV coat antigens.

Adult↗

PubMed1984

Immunogenic and antigenic epitopes of immunoglobulins--VII. The topographical distribution of Fc gamma epitopes and the relationship of an iso-allotypic specificity to the presence of histidine 435.

Epitopes recognised by a panel of 23 anti-Fc gamma monoclonal antibodies (McAbs) have been subdivided into three groups each having a distinct topographical distribution. One group of mutually inhibitory McAbs are reactive with epitopes expressed on the fy "surface" of the C gamma 2 domain. A second group recognises epitopes in the region of arginine 355 of the C gamma 3 domain whilst the third group recognises epitopes expressed in the inter C gamma 2/C gamma 3 domain region--as evidenced by inhibition of Staphylococcus aureus protein A binding. An antibody of the latter group reactive with IgG1, 2, 4 and IgG3m(15,16) proteins but not IgG3m(5) or IgG3m(21) proteins allows histidine 435 to be identified as a critical residue for expression of the epitope recognised by this antibody.

Animals↗

PubMed1984

T-cell proliferation and expression of MHC class II antigens.

Two monoclonal antibodies to major histocompatibility complex (MHC) class II antigens, which in combination identify beta chains encoded by the SB and DR loci, were used to investigate which of these gene products were expressed at the cell surface of unstimulated T cells and at various stages of mitogen-induced T-cell maturation. In tests on blood lymphocytes from healthy donors 12% of T cells expressed class II antigens, but only SB antigens were expressed. During activation of T lymphocytes, SB-coded antigens were expressed before DR antigens, and the kinetics of SB expression correlated with the proliferative response of T cells. These results and consideration of recent reports from other laboratories lead us to suggest that SB-coded class II antigens play a role in T-cell proliferation.

Cells, Cultured↗

PubMed1984

Monoclonal antibodies to an immunoglobulin allotype marker G1m(f).

The properties of a group of mouse monoclonal antibodies (McAbs) specific for the human G1m(f) allotype marker on immunoglobulin G are described. The specificity of all 5 McAbs was anti-Gm(f) in haemagglutination assays detecting the G1m(f) determinant on 6 ng of purified G1m(f) paraproteins. A high dilution (greater than or equal to 1/10(4)) could be used for the majority of McAbs in this assay. In Elisa the G1m(f) marker could be detected in homozygote G1m(f+z-) sera at a serum dilution of greater than or equal to 1/10(4). In Elisa assays the G1m(f) specificity was lost when IgG was bound directly to polystyrene but was restored when IgG was bound via anti-human IgG to the polystyrene plate. A possible conformational change in IgG to account for this loss of specificity is discussed. It is expected that these McAbs with their high titre and increased sensitivity over conventional Gm antisera will allow more detailed analysis of the Gm marker system.

Animals↗

PubMed1984

Investigation of the idiotypic determinants of IgM monoclonal proteins associated with neuropathy.

Occasionally IgM monoclonal proteins (MP) from benign gammopathies bind neural antigens and have been associated with demyelinating peripheral neuropathy. A monoclonal antibody (McAb) has been raised against a cross-reactive idiotypic determinant expressed by three out of 12 MP with specificity for neural antigens. The determinant recognised by this MoAb is only expressed when both H and L chain are properly associated. The idiotypic determinant was not expressed by 130 MP randomly selected from patients without neuropathy nor in polyclonal immunoglobulin preparations obtained from healthy individuals.

Antibodies, Monoclonal↗